Judit Kucsera

Evolution of linear chromosomes and multipartite genomes in yeast mitochondria

Mitochondrial genome diversity in closely related species provides an excellent platform for investigation of chromosome architecture and its evolution by means of comparative genomics. In this study, we determined the complete mitochondrial DNA sequences of eight Candida species and analyzed their molecular architectures. Our survey revealed a puzzling variability of genome architecture, including circular- and linear-mapping and multipartite linear forms. We propose that the arrangement of large inverted repeats identified in these genomes plays a crucial role in alterations of their molecular architectures. In specific arrangements, the inverted repeats appear to function as resolution elements, allowing genome conversion among different topologies, eventually leading to genome fragmentation into multiple linear DNA molecules. We suggest that molecular transactions generating linear mitochondrial DNA molecules with defined telomeric structures may parallel the evolutionary emergence of linear chromosomes and multipartite genomes in general and may provide clues for the origin of telomeres and pathways implicated in their maintenance.

Homothallic life cycle in the diploid red yeast Xanthophyllomyces dendrorhous (Phaffia rhodozyma)

Sexual activity was induced in the basidiomyceteous Phaffia rhodozyma (Xanthophyllomyces dendrorhous) by depletion of nitrogen from the culture medium. This activity involved both mating between two yeast cells and the formation of basidiospores. Mating is possibly started by a G1 phase arrest of the cell cycle, as in other yeasts. The life cycle exhibited homothallic features. Crosses between genetically marked strains, and pulse-field gel electrophoresis of the chromosomal DNA of cells derived from individual spores revealed evidence of karyogamy, meiosis and even recombination. The segregation ratio in tetrads pointed to diploid vegetative cells, which formed tetraploid zygotes and the immediate meiosis then gave rise to diploid progenies again. Apart from the type strain Phaffia rhodozyma CBS 5905, all the examined strains were able to sporulate.

Variability and inheritance of double stranded RNA viruses in Phaffia rhodozyma

Abstract The present survey demonstrates polymorphism in both the length and the number of double-stranded RNAs (dsRNAs) among six Phaffia rhodozyma strains. Strains with one-, three- and four-types of dsRNA molecules were found, while two strains proved to be dsRNA-free. Elongated icosahedral virus-like particles (VLPs) 34×26 nm in size were detected in strains carrying four- or three-types of dsRNAs. One 3.7-kb dsRNA molecule was found not to form part of the VLP genome. Transmission of the VLPs of strain ATCC 24203 was followed through the basidiospores during the sexual cycle. Cytoplasmic inheritance was observed.

The mitochondrial genome of the pathogenic yeast Candida subhashii: GC-rich linear DNA with a protein covalently attached to the 5' termini.

As a part of our initiative aimed at a large-scale comparative analysis of fungal mitochondrial genomes, we determined the complete DNA sequence of the mitochondrial genome of the yeast Candida subhashii and found that it exhibits a number of peculiar features. First, the mitochondrial genome is represented by linear dsDNA molecules of uniform length (29 795 bp), with an unusually high content of guanine and cytosine residues (52.7 %). Second, the coding sequences lack introns; thus, the genome has a relatively compact organization. Third, the termini of the linear molecules consist of long inverted repeats and seem to contain a protein covalently bound to terminal nucleotides at the 5′ ends. This architecture resembles the telomeres in a number of linear viral and plasmid DNA genomes classified as invertrons, in which the terminal proteins serve as specific primers for the initiation of DNA synthesis. Finally, although the mitochondrial genome of C. subhashii contains essentially the same set of genes as other closely related pathogenic Candida species, we identified additional ORFs encoding two homologues of the family B protein-priming DNA polymerases and an unknown protein. The terminal structures and the genes for DNA polymerases are reminiscent of linear mitochondrial plasmids, indicating that this genome architecture might have emerged from fortuitous recombination between an ancestral, presumably circular, mitochondrial genome and an invertron-like element.

Hybridization Studies by Crossing and Protoplast Fusion within the Genus Schizosaccharomyces Lindner

Summary: Hybridization studies based on the use of crossing and the protoplast fusion technique revealed the relationship between the various taxa of the yeast genus Schizosaccharomyces Lindner. From a series of intra- and interspecific crosses as well as protoplast fusions, we concluded that a revision of the genus is required. Being interfertile, S. pombe and S. malidevorans have to be included in one species S. pombe. From the protoplast fusion results, this new taxon seems to be closely related to S. octosporus. The results obtained are discussed in connection with the characteristics of the low temperature spectra of the cytochromes.

Differences in mitochondrial genome organization of Cryptococcus neoformans strains.

The organization of the mitochondrial genomes in two strains belonging in different varieties of Cryptococcus neoformans was analysed. Physical maps of the mtDNA of the IFM5844 (var. neoformans) and IFO410 (var. grubii) strains were constructed by using EcoRI and EcoRV restriction enzymes; functional maps were constructed by hybridization, cloning and sequencing. Most of the genes important in the mitochondrial function (ND1, ND2, ND3, ND4, ND4L, ND5, ND6, ATP6, ATP9, COX1, COX2 and COB) and protein synthesis (SsrRNA and LsrRNA) were localized. We did not find any differences between the strains in the order of these genes. However, they differed significantly in the sizes of the mtDNAs: 32.6 kb for IFM5844, and 24.1 kb for IFO410. This can be attributed to two large regions of the mtDNA. In these regions, differences were found in the numbers of introns in COX1 (no intron in var. grubii, 5 introns in var. neoformans), COB (1 intron in var. grubii, 2 introns in var. neoformans), LsrRNA (no intron in var. grubii, 2 introns in var. neoformans), and ND5 (no intron in var. grubii, 1 intron in var. neoformans) genes. In several introns of the COB and COX1 genes LAGLIDADG motifs were found. Differences were also observed in the nucleotide sequences of some genes and in the sizes and sequences of intergenic regions. The nucleotide sequences of the genes of the IFM and IFO strains were compared with those of the H-99 and JEC 21 strains from the database. Surprisingly high similarities were found between the strains belonging in var. grubii (IFO 410 and H-99) and var. neoformans (IFM 5844 and JEC 21).

Comparative analysis of the complete mitochondrial genomes of Aspergillus niger mtDNA type 1a and Aspergillus tubingensis mtDNA type 2b.

To understand the differences in the organization of mitochondrial genomes of the very closely related Aspergillus niger and Aspergillus tubingensis species, we determined the complete genome sequence of the 1a mtDNA type of A. niger and 2b mtDNA type of A. tubingensis and now we provide a comparative analysis of the two mtDNAs. We found that (1) the organization (gene content and order) of the two genomes is almost identical and (2) the size difference between them is principally attributed to the different intron content of their cox1, atp9 and ndh4L genes.

Mycocin production in Cryptococcus aquaticus

Double-stranded RNA viruses of about 35 nm in diameter were isolated from a mycocin-secreting strain of Cryptococcus aquaticus. A derivative of this strain, lacking small dsRNA, was non-mycocinogenic and sensitive to its own toxin. The killing pattern of this mycocin was restricted to some species of the Cystofilobasidiales clade. Despite the differences in genome size of dsRNA viruses in mycocinogenic strains of Cryptococcus aquaticus, Cystofilobasidium sp. CBS 6569, Cystofilobasidium bisporidii, Cystofilobasidium infirmominiatum, Trichosporon pullulans and Xanthophyllomyces dendrorhous and killing patterns of their mycocins, the viral genomes showed homology in hybridisation experiments.

Interpretation of mtDNA RFLP variability among Aspergillus tubingensis isolates

Aspergillus tubingensis isolates collected from distant geographic areas were earlier classified into six groups on the basis of the mtDNA RFLP variability they exhibited (mtDNA types 2a–2f). In the present work, we investigated the reason for the intraspecific mtDNA variability and we describe here how this fungus, with a relatively small mitochondrial genome, can display intraspecific polymorphism due to intron acquisition and also sporadic point mutations affecting the recognition motifs of the restriction enzymes employed in the RFLP analysis. Three different LAGLI-DADG type group I introns were identified in the cox1 gene amongst the six mtDNA RFLP types. MtDNAs of types 2b and 2d contain all of the three introns, mtDNA of type 2f carries only one, and the other mtDNA types contain two introns each. Comparative analysis showed that the first and second introns of mtDNAs of types 2b and 2d are well distributed among fungi, indicating their active horizontal transfer capacity. The third intron occurs rarely among fungi and is restricted to a limited number of fungal species, namely to A. tubingensis and the yeast Candida stellata. It is interesting that this intron is present in a small mitochondrial genome such as that of A. tubingensis and, considering its rarity, its presence amongst black Aspergillus isolates is recommended to be considered as a tool to establish taxonomical unit(s) or to track down evolutionary divergence of closely related taxonomical units.

Biology of the red yeast Xanthophyllomyces dendrorhous (Phaffia rhodozyma)

Phaffia rhodozyma is a red-pigmented basidiomycetous yeast that produces astaxanthin. Because of this property, it is fermented commercially on a large scale. Astaxanthin is used as a food colorant for fish, and as an antioxidant, it is potentially useful in the human pharmaceutical industry. This review summarizes the published biology of this yeast: its morphology and ultrastructure, organization of the cytoskeleton, and the nuclear and extrachromosomal genomes. Alteration of sexual and vegetative phases in the life cycle and its biotechnological importance are also described.