Introducing a new breed of wine yeast: interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast and Saccharomyces mikatae

Interspecific hybrids are commonplace in agriculture and horticulture; bread wheat and grapefruit are but two examples. The benefits derived from interspecific hybridisation include the potential of generating advantageous transgressive phenotypes. This paper describes the generation of a new breed of wine yeast by interspecific hybridisation between a commercial Saccharomyces cerevisiae wine yeast strain and Saccharomyces mikatae, a species hitherto not associated with industrial fermentation environs. While commercially available wine yeast strains provide consistent and reliable fermentations, wines produced using single inocula are thought to lack the sensory complexity and rounded palate structure obtained from spontaneous fermentations. In contrast, interspecific yeast hybrids have the potential to deliver increased complexity to wine sensory properties and alternative wine styles through the formation of novel, and wider ranging, yeast volatile fermentation metabolite profiles, whilst maintaining the robustness of the wine yeast parent. Screening of newly generated hybrids from a cross between a S. cerevisiae wine yeast and S. mikatae (closely-related but ecologically distant members of the Saccharomyces sensu stricto clade), has identified progeny with robust fermentation properties and winemaking potential. Chemical analysis showed that, relative to the S. cerevisiae wine yeast parent, hybrids produced wines with different concentrations of volatile metabolites that are known to contribute to wine flavour and aroma, including flavour compounds associated with non-Saccharomyces species. The new S. cerevisiae x S. mikatae hybrids have the potential to produce complex wines akin to products of spontaneous fermentation while giving winemakers the safeguard of an inoculated ferment.Jennifer R. Bellon, Frank Schmid, Dimitra L. Capone, Barbara L. Dunn, Paul J. Chamber

Systems-based approaches enable identification of gene targets which improve the flavour profile of low-ethanol wine yeast strains

Metabolic engineering has been vital to the development of industrial microbes such as the yeast Saccharomyces cerevisiae. However, sequential rounds of modification are often needed to achieve particular industrial design targets. Systems biology approaches can aid in identifying genetic targets for modification through providing an integrated view of cellular physiology. Recently, research into the generation of commercial yeasts that can produce reduced-ethanol wines has resulted in metabolically-engineered strains of S. cerevisiae that are less efficient at producing ethanol from sugar. However, these modifications led to the concomitant production of off-flavour by-products. A combination of transcriptomics, proteomics and metabolomics was therefore used to investigate the physiological changes occurring in an engineered low-ethanol yeast strain during alcoholic fermentation. Integration of ‘omics data identified several metabolic reactions, including those related to the pyruvate node and redox homeostasis, as being significantly affected by the low-ethanol engineering methodology, and highlighted acetaldehyde and 2,4,5-trimethyl-1,3-dioxolane as the main off-flavour compounds. Gene remediation strategies were then successfully applied to decrease the formation of these by-products, while maintaining the ‘low-alcohol’ phenotype. The data generated from this comprehensive systems-based study will inform wine yeast strain development programmes, which, in turn, could potentially play an important role in assisting winemakers in their endeavour to produce low-alcohol wines with desirable flavour profiles

Insights into the Dekkera bruxellensis genomic landscape: comparative genomics reveals variations in ploidy and nutrient utilisation potential amongst wine isolates

The yeast Dekkera bruxellensis is a major contaminant of industrial fermentations, such as those used for the production of biofuel and wine, where it outlasts and, under some conditions, outcompetes the major industrial yeast Saccharomyces cerevisiae. In order to investigate the level of inter-strain variation that is present within this economically important species, the genomes of four diverse D. bruxellensis isolates were compared. While each of the four strains was shown to contain a core diploid genome, which is clearly sufficient for survival, two of the four isolates have a third haploid complement of chromosomes. The sequences of these additional haploid genomes were both highly divergent from those comprising the diploid core and divergent between the two triploid strains. Similar to examples in the Saccharomyces spp. clade, where some allotriploids have arisen on the basis of enhanced ability to survive a range of environmental conditions, it is likely these strains are products of two independent hybridisation events that may have involved multiple species or distinct sub-species of Dekkera. Interestingly these triploid strains represent the vast majority (92%) of isolates from across the Australian wine industry, suggesting that the additional set of chromosomes may confer a selective advantage in winery environments that has resulted in these hybrid strains all-but replacing their diploid counterparts in Australian winery settings. In addition to the apparent inter-specific hybridisation events, chromosomal aberrations such as strain-specific insertions and deletions and loss-of-heterozygosity by gene conversion were also commonplace. While these events are likely to have affected many phenotypes across these strains, we have been able to link a specific deletion to the inability to utilise nitrate by some strains of D. bruxellensis, a phenotype that may have direct impacts in the ability for these strains to compete with S. cerevisiae.Anthony R. Borneman, Ryan Zeppel, Paul J. Chambers, Chris D. Curti

Functional Divergence in the Genus Oenococcus as Predicted by Genome Sequencing of the Newly-Described Species, Oenococcus kitaharae

Oenococcus kitaharae is only the second member of the genus Oenococcus to be identified and is the closest relative of the industrially important wine bacterium Oenococcus oeni. To provide insight into this new species, the genome of the type strain of O. kitaharae, DSM 17330, was sequenced. Comparison of the sequenced genomes of both species show that the genome of O. kitaharae DSM 17330 contains many genes with predicted functions in cellular defence (bacteriocins, antimicrobials, restriction-modification systems and a CRISPR locus) which are lacking in O. oeni. The two genomes also appear to differentially encode several metabolic pathways associated with amino acid biosynthesis and carbohydrate utilization and which have direct phenotypic consequences. This would indicate that the two species have evolved different survival techniques to suit their particular environmental niches. O. oeni has adapted to survive in the harsh, but predictable, environment of wine that provides very few competitive species. However O. kitaharae appears to have adapted to a growth environment in which biological competition provides a significant selective pressure by accumulating biological defence molecules, such as bacteriocins and restriction-modification systems, throughout its genome

Circular DNA Intermediate in the Duplication of Nile Tilapia vasa Genes

vasa is a highly conserved RNA helicase involved in animal germ cell development. Among vertebrate species, it is typically present as a single copy per genome. Here we report the isolation and sequencing of BAC clones for Nile tilapia vasa genes. Contrary to a previous report that Nile tilapia have a single copy of the vasa gene, we find evidence for at least three vasa gene loci. The vasa gene locus was duplicated from the original site and integrated into two distant novel sites. For one of these insertions we find evidence that the duplication was mediated by a circular DNA intermediate. This mechanism of gene duplication may explain the origin of isolated gene duplicates during the evolution of fish genomes. These data provide a foundation for studying the role of multiple vasa genes in the development of tilapia gonads, and will contribute to investigations of the molecular mechanisms of sex determination and evolution in cichlid fishes

Prediction of Phenotype-Associated Genes via a Cellular Network Approach: A Candida albicans Infection Case Study

Candida albicans is the most prevalent opportunistic fungal pathogen in humans causing superficial and serious systemic infections. The infection process can be divided into three stages: adhesion, invasion, and host cell damage. To enhance our understanding of these C. albicans infection stages, this study aimed to predict phenotype-associated genes involved during these three infection stages and their roles in C. albicans–host interactions. In light of the principles that proteins that lie closer to one another in a protein interaction network are more likely to have similar functions, and that genes regulated by the same transcription factors tend to have similar functions, a cellular network approach was proposed to predict the phenotype-associated genes in this study. A total of 4, 12, and 3 genes were predicted as adhesion-, invasion-, and damage-associated genes during C. albicans infection, respectively. These predicted genes highlight the facts that cell surface components are critical for cell adhesion, and that morphogenesis is crucial for cell invasion. In addition, they provide targets for further investigations into the mechanisms of the three C. albicans infection stages. These results give insights into the responses elicited in C. albicans during interaction with the host, possibly instrumental in identifying novel therapies to treat C. albicans infection

Conservation and divergence of known apicomplexan transcriptional regulons

<p>Abstract</p> <p>Background</p> <p>The apicomplexans are a diverse phylum of parasites causing an assortment of diseases including malaria in a wide variety of animals and lymphoproliferation in cattle. Little is known about how these varied parasites regulate their transcriptional regulons. Even less is known about how regulon systems, consisting of transcription factors and target genes together with their associated biological process, evolve in these diverse parasites.</p> <p>Results</p> <p>In order to obtain insights into the differences in transcriptional regulation between these parasites we compared the orthology profiles of putative malaria transcription factors across species and examined the enrichment patterns of four binding sites across eleven apicomplexans.</p> <p>About three-fifths of the factors are broadly conserved in several phylogenetic orders of sequenced apicomplexans. This observation suggests the existence of regulons whose regulation is conserved across this ancient phylum. Transcription factors not broadly conserved across the phylum are possibly involved in regulon systems that have diverged between species. Examining binding site enrichment patterns in light of transcription factor conservation patterns suggests a second mode via which regulon systems may diverge - rewiring of existing transcription factors and their associated binding sites in specific ways. Integrating binding sites with transcription factor conservation patterns also facilitated prediction of putative regulators for one of the binding sites.</p> <p>Conclusions</p> <p>Even though transcription factors are underrepresented in apicomplexans, the distribution of these factors and their associated regulons reflect common and family-specific transcriptional regulatory processes.</p

BayesPI - a new model to study protein-DNA interactions: a case study of condition-specific protein binding parameters for Yeast transcription factors

<p>Abstract</p> <p>Background</p> <p>We have incorporated Bayesian model regularization with biophysical modeling of protein-DNA interactions, and of genome-wide nucleosome positioning to study protein-DNA interactions, using a high-throughput dataset. The newly developed method (BayesPI) includes the estimation of a transcription factor (TF) binding energy matrices, the computation of binding affinity of a TF target site and the corresponding chemical potential.</p> <p>Results</p> <p>The method was successfully tested on synthetic ChIP-chip datasets, real yeast ChIP-chip experiments. Subsequently, it was used to estimate condition-specific and species-specific protein-DNA interaction for several yeast TFs.</p> <p>Conclusion</p> <p>The results revealed that the modification of the protein binding parameters and the variation of the individual nucleotide affinity in either recognition or flanking sequences occurred under different stresses and in different species. The findings suggest that such modifications may be adaptive and play roles in the formation of the environment-specific binding patterns of yeast TFs and in the divergence of TF binding sites across the related yeast species.</p

The Stress Response Factors Yap6, Cin5, Phd1, and Skn7 Direct Targeting of the Conserved Co-Repressor Tup1-Ssn6 in S. cerevisiae

Maintaining the proper expression of the transcriptome during development or in response to a changing environment requires a delicate balance between transcriptional regulators with activating and repressing functions. The budding yeast transcriptional co-repressor Tup1-Ssn6 is a model for studying similar repressor complexes in multicellular eukaryotes. Tup1-Ssn6 does not bind DNA directly, but is directed to individual promoters by one or more DNA-binding proteins, referred to as Tup1 recruiters. This functional architecture allows the Tup1-Ssn6 to modulate the expression of genes required for the response to a variety of cellular stresses. To understand the targeting or the Tup1-Ssn6 complex, we determined the genomic distribution of Tup1 and Ssn6 by ChIP-chip. We found that most loci bound by Tup1-Ssn6 could not be explained by co-occupancy with a known recruiting cofactor and that deletion of individual known Tup1 recruiters did not significantly alter the Tup1 binding profile. These observations suggest that new Tup1 recruiting proteins remain to be discovered and that Tup1 recruitment typically depends on multiple recruiting cofactors. To identify new recruiting proteins, we computationally screened for factors with binding patterns similar to the observed Tup1-Ssn6 genomic distribution. Four top candidates, Cin5, Skn7, Phd1, and Yap6, all known to be associated with stress response gene regulation, were experimentally confirmed to physically interact with Tup1 and/or Ssn6. Incorporating these new recruitment cofactors with previously characterized cofactors now explains the majority of Tup1 targeting across the genome, and expands our understanding of the mechanism by which Tup1-Ssn6 is directed to its targets