Silvia Poláková

Formation of new chromosomes as a virulence mechanism in yeast Candida glabrata.

In eukaryotes, the number and rough organization of chromosomes is well preserved within isolates of the same species. Novel chromosomes and loss of chromosomes are infrequent and usually associated with pathological events. Here, we analyzed 40 pathogenic isolates of a haploid and asexual yeast, Candida glabrata, for their genome structure and stability. This organism has recently become the second most prevalent yeast pathogen in humans. Although the gene sequences were well conserved among different strains, their chromosome structures differed drastically. The most frequent events reshaping chromosomes were translocations of chromosomal arms. However, also larger segmental duplications were frequent and occasionally we observed novel chromosomes. Apparently, this yeast can generate a new chromosome by duplication of chromosome segments carrying a centromere and subsequently adding novel telomeric ends. We show that the observed genome plasticity is connected with antifungal drug resistance and it is likely an advantage in the human body, where environmental conditions fluctuate a lot.

A Second Pathway to Degrade Pyrimidine Nucleic Acid Precursors in Eukaryotes.

Pyrimidine bases are the central precursors for RNA and DNA, and their intracellular pools are determined by de novo, salvage and catabolic pathways. In eukaryotes, degradation of uracil has been believed to proceed only via the reduction to dihydrouracil. Using a yeast model, Saccharomyces kluyveri, we show that during degradation, uracil is not reduced to dihydrouracil. Six loci, named URC1-6 (for uracil catabolism), are involved in the novel catabolic pathway. Four of them, URC3,5, URC6, and URC2 encode urea amidolyase, uracil phosphoribosyltransferase, and a putative transcription factor, respectively. The gene products of URC1 and URC4 are highly conserved proteins with so far unknown functions and they are present in a variety of prokaryotes and fungi. In bacteria and in some fungi, URC1 and URC4 are linked on the genome together with the gene for uracil phosphoribosyltransferase (URC6). Urc1p and Urc4p are therefore likely the core components of this novel biochemical pathway. A combination of genetic and analytical chemistry methods demonstrates that uridine monophosphate and urea are intermediates, and 3-hydroxypropionic acid, ammonia and carbon dioxide the final products of degradation. The URC pathway does not require the presence of an active respiratory chain and is therefore different from the oxidative and rut pathways described in prokaryotes, although the latter also gives 3-hydroxypropionic acid as the end product. The genes of the URC pathway are not homologous to any of the eukaryotic or prokaryotic genes involved in pyrimidine degradation described to date.

Mitochondrial genome from the facultative anaerobe and petite‐positive yeast Dekkera bruxellensis contains the NADH dehydrogenase subunit genes

The progenitor of the Dekkera/Brettanomyces clade separated from the Saccharomyces/Kluyveromyces clade over 200 million years ago. However, within both clades, several lineages developed similar physiological traits. Both Saccharomyces cerevisiae and Dekkera bruxellensis are facultative anaerobes; in the presence of excess oxygen and sugars, they accumulate ethanol (Crabtree effect) and they both spontaneously generate respiratory-deficient mutants (petites). In order to understand the role of respiratory metabolism, the mitochondrial DNA (mtDNA) molecules of two Dekkera/Brettanomyces species were analysed. Dekkera bruxellensis mtDNA shares several properties with S. cerevisiae, such as the large genome size (76 453 bp), and the organization of the intergenic sequences consisting of spacious AT-rich regions containing a number of hairpin GC-rich cluster-like elements. In addition to a basic set of the mitochondrial genes coding for the components of cytochrome oxidase, cytochrome b, subunits of ATPase, two rRNA subunits and 25 tRNAs, D. bruxellensis also carries genes for the NADH dehydrogenase complex. Apparently, in yeast, the loss of this complex is not a precondition to develop a petite-positive, Crabtree-positive and anaerobic nature. On the other hand, mtDNA from a petite-negative Brettanomyces custersianus is much smaller (30 058 bp); it contains a similar gene set and has only short intergenic sequences.

Geotrichum bryndzae sp. nov., a novel asexual arthroconidial yeast species related to the genus Galactomyces

Ten strains of an asexual arthroconidial yeast species were isolated from Bryndza, a traditional Slovak artisanal sheep cheese, which was manufactured from raw milk during a 4-month summer production period at two Slovakian sites (the northern RuZomberok and the central-southern Tisovec areas). Sequence comparison of the D1/D2 domains of the large-subunit rRNA gene revealed that this yeast represents a novel species of the genus Geotrichum, which contains anamorphs of the ascogenous genus Galactomyces, for which the name Geotrichum bryndzae sp. nov. is proposed (type culture CCY 16-2-1T=NRRL Y-48450T=CBS 11176T). The novel species is most closely related to Geotrichum silvicola NRRL Y-27641T, although yeasts with identical or very similar sequences have been found throughout the world.

A complete sequence of Saccharomyces paradoxus mitochondrial genome that restores the respiration in S. cerevisiae

We determined the complete sequence of 71 355-bp-long mitochondrial genome from Saccharomyces paradoxus entirely by direct sequencing of purified mitochondrial DNA (mtDNA). This mtDNA possesses the same features as its close relative Saccharomyces cerevisiae - A + T content 85.9%, set of genes coding for the three components of cytochrome oxidase, cytochrome b, three subunits of ATPase, both ribosomal subunits, gene for ribosomal protein, rnpB gene, tRNA package (24) and yeast genetic code. Genes are interrupted by nine group I and group II introns, two of which are in positions unknown in S. cerevisiae, but recognized in Saccharomyces pastorianus. The gene products are related to S. cerevisiae, and the identity of amino acid residues varies from 100% for cox2 to 83% for rps3. The remarkable differences from S. cerevisiae are (1) different gene order (translocation of trnF-trnT1-trnV-cox3-trnfM-rnpb-trnP and transposition of trnW-rns), (2) occurrence of two unusual GI introns, (3) eight active ori elements, and (4) reduced number of GC clusters and divergent intergenic spacers. Despite these facts, the sequenced S. paradoxus mtDNA introduced to S. cerevisiae was able to support the respiratory function to the same extent as the original mtDNAs.

Post-zygotic sterility and cytonuclear compatibility limits in S. cerevisiae xenomitochondrial cybrids

Nucleo-mitochondrial interactions, particularly those determining the primary divergence of biological species, can be studied by means of xenomitochondrial cybrids, which are cells where the original mitochondria are substituted by their counterparts from related species. Saccharomyces cerevisiae cybrids are prepared simply by the mating of the ρ0 strain with impaired karyogamy and germinating spores from other Saccharomyces species and fall into three categories. Cybrids with compatible mitochondrial DNA (mtDNA) from Saccharomyces paradoxus CBS 432 and Saccharomyces cariocanus CBS 7994 are metabolically and genetically similar to cybrids containing mtDNA from various S. cerevisiae. Cybrids with mtDNA from other S. paradoxus strains, S. cariocanus, Saccharomyces kudriavzevii, and Saccharomyces mikatae require a period of adaptation to establish efficient oxidative phosphorylation. They exhibit a temperature-sensitive phenotype, slower growth rate on a non-fermentable carbon source and a long lag phase after the shift from glucose. Their decreased respiration capacity and reduced cytochrome aa3 content is associated with the inefficient splicing of cox1I3β, the intron found in all Saccharomyces species but not in S. cerevisiae. The splicing defect is compensated in cybrids by nuclear gain-of-function and can be alternatively suppressed by overexpression of MRP13 gene for mitochondrial ribosomal protein or the MRS2, MRS3, and MRS4 genes involved in intron splicing. S. cerevisiae with Saccharomyces bayanus mtDNA is unable to respire and the growth on ethanol–glycerol can be restored only after mating to some mit− strains. The nucleo-mitochondrial compatibility limit of S. cerevisiae and other Saccharomyces was set between S. kudriavzevii and S. bayanus at the divergence from S. cerevisiae about 15 MYA. The MRS1-cox1 S. cerevisiae/S. paradoxus cytonuclear Dobzhansky–Muller pair has a neglible impact on the separation of species since its imperfection is compensated for by gain-of-function mutation.

The evolutionary history of Saccharomyces species inferred from completed mitochondrial genomes and revision in the ‘yeast mitochondrial genetic code’

Abstract The yeast Saccharomyces are widely used to test ecological and evolutionary hypotheses. A large number of nuclear genomic DNA sequences are available, but mitochondrial genomic data are insufficient. We completed mitochondrial DNA (mtDNA) sequencing from Illumina MiSeq reads for all Saccharomyces species. All are circularly mapped molecules decreasing in size with phylogenetic distance from Saccharomyces cerevisiae but with similar gene content including regulatory and selfish elements like origins of replication, introns, free-standing open reading frames or GC clusters. Their most profound feature is species-specific alteration in gene order. The genetic code slightly differs from well-established yeast mitochondrial code as GUG is used rarely as the translation start and CGA and CGC code for arginine. The multilocus phylogeny, inferred from mtDNA, does not correlate with the trees derived from nuclear genes. mtDNA data demonstrate that Saccharomyces cariocanus should be assigned as a separate species and Saccharomyces bayanus CBS 380T should not be considered as a distinct species due to mtDNA nearly identical to Saccharomyces uvarum mtDNA. Apparently, comparison of mtDNAs should not be neglected in genomic studies as it is an important tool to understand the origin and evolutionary history of some yeast species.

Erratum to: Small chromosomes among Danish Candida glabrata isolates originated through different mechanisms.

The online version of the original article can be found under doi:10.1007/s10482-013-9931-3.