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Yeast Genetics 2021

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A special issue of Journal of Fungi (ISSN 2309-608X). This special issue belongs to the section "Fungal Genomics, Genetics and Molecular Biology".

Deadline for manuscript submissions: closed (30 November 2021) | Viewed by 27952

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Dr. Ivan-Kresimir Svetec

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Department of Biochemical Engineering, Faculty of Food Technology and Biotechnology, University of Zagreb, Zagreb, Croatia
Interests: yeast molecular genetics; genetic recombination; DNA repair; genome stability
Special Issues, Collections and Topics in MDPI journals

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Dear Colleagues,

Yeast Saccharomyces cerevisiae is not only one of the most important industrial microorganisms but also a model organism for investigation of almost all biological processes in eukaryotes. This conventional yeast was the first eukariote to be successfully transformed with exogenous DNA. Due to efficient homologous recombination between transforming DNA and its genome, it is easy to precisely genetically modify this yeast, allowing the construction of both biotechnologically relevant strains and strains used in scientific research. On the other hand, there is a huge number of uncharacterized or poorly characterized nonconventional yeasts, and it is already known that at least some of them have certain advantages over conventional yeasts, such as efficient use of alternative carbon sources, higher resistance to growth and fermentation inhibitors, production of interesting metabolites, etc. However, in a great majority of these nonconventional yeasts, the construction of desired strains is not a routine procedure because molecular tools for precise modification of their genome have not been developed yet. Therefore, the focus of this Special Issue is on research on the genetics of conventional and nonconventional yeast, as well as development of molecular tools for targeted modification of yeasts genomes and construction of potentially biotechnologically relevant yeast strains.

Dr. Ivan-Kresimir Svetec
Guest Editor

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Keywords

Saccharomyces cerevisiae
conventional yeasts
nonconventional yeasts
genetics
genome stability
genetic recombination
DNA repair
gene expression
strain construction
genetic transformation
targeted genetic modification

Related Special Issue

Yeast Genetics 2022 in Journal of Fungi (5 articles)

Published Papers (11 papers)

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Research

Jump to: Review

15 pages, 5298 KiB

Open AccessArticle

Toolbox for Genetic Transformation of Non-Conventional Saccharomycotina Yeasts: High Efficiency Transformation of Yeasts Belonging to the Schwanniomyces Genus

by Angela Matanović, Kristian Arambašić, Bojan Žunar, Anamarija Štafa, Marina Svetec Miklenić, Božidar Šantek and Ivan-Krešimir Svetec

J. Fungi 2022, 8(5), 531; https://0-doi-org.brum.beds.ac.uk/10.3390/jof8050531 - 20 May 2022

Viewed by 2222

Abstract

Non-conventional yeasts are increasingly being investigated and used as producers in biotechnological processes which often offer advantages in comparison to traditional and well-established systems. Most biotechnologically interesting non-conventional yeasts belong to the Saccharomycotina subphylum, including those already in use (Pichia pastoris, Yarrowia lypolitica, etc.), as well as those that are promising but as yet insufficiently characterized. Moreover, for many of these yeasts the basic tools of genetic engineering needed for strain construction, including a procedure for efficient genetic transformation, heterologous protein expression and precise genetic modification, are lacking. The first aim of this study was to construct a set of integrative and replicative plasmids which can be used in various yeasts across the Saccharomycotina subphylum. Additionally, we demonstrate here that the electroporation procedure we developed earlier for transformation of B. bruxellensis can be applied in various yeasts which, together with the constructed plasmids, makes a solid starting point when approaching a transformation of yeasts form the Saccharomycotina subphylum. To provide a proof of principle, we successfully transformed three species from the Schwanniomyces genus (S. polymorphus var. polymorphus, S. polymorphus var. africanus and S. pseudopolymorphus) with high efficiencies (up to 8 × 10³ in case of illegitimate integration of non-homologous linear DNA and up to 4.7 × 10⁵ in case of replicative plasmid). For the latter two species this is the first reported genetic transformation. Moreover, we found that a plasmid carrying replication origin from Scheffersomyces stipitis can be used as a replicative plasmid for these three Schwanniomyces species. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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19 pages, 3727 KiB

Open AccessArticle

Near Chromosome-Level Genome Assembly and Annotation of Rhodotorula babjevae Strains Reveals High Intraspecific Divergence

by Giselle C. Martín-Hernández, Bettina Müller, Christian Brandt, Martin Hölzer, Adrian Viehweger and Volkmar Passoth

J. Fungi 2022, 8(4), 323; https://0-doi-org.brum.beds.ac.uk/10.3390/jof8040323 - 22 Mar 2022

Cited by 1 | Viewed by 2386

Abstract

The genus Rhodotorula includes basidiomycetous oleaginous yeast species. Rhodotorula babjevae can produce compounds of biotechnological interest such as lipids, carotenoids, and biosurfactants from low value substrates such as lignocellulose hydrolysate. High-quality genome assemblies are needed to develop genetic tools and to understand fungal evolution and genetics. Here, we combined short- and long-read sequencing to resolve the genomes of two R. babjevae strains, CBS 7808 (type strain) and DBVPG 8058, at chromosomal level. Both genomes are 21 Mbp in size and have a GC content of 68.2%. Allele frequency analysis indicates that both strains are tetraploid. The genomes consist of a maximum of 21 chromosomes with a size of 0.4 to 2.4 Mbp. In both assemblies, the mitochondrial genome was recovered in a single contig, that shared 97% pairwise identity. Pairwise identity between most chromosomes ranges from 82 to 87%. We also found indications for strain-specific extrachromosomal endogenous DNA. A total of 7591 and 7481 protein-coding genes were annotated in CBS 7808 and DBVPG 8058, respectively. CBS 7808 accumulated a higher number of tandem duplications than DBVPG 8058. We identified large translocation events between putative chromosomes. Genome divergence values between the two strains indicate that they may belong to different species. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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18 pages, 2405 KiB

Open AccessArticle

Purine Auxotrophic Starvation Evokes Phenotype Similar to Stationary Phase Cells in Budding Yeast

by Agnese Kokina, Kristel Tanilas, Zane Ozolina, Karlis Pleiko, Karlis Shvirksts, Ilze Vamza and Janis Liepins

J. Fungi 2022, 8(1), 29; https://0-doi-org.brum.beds.ac.uk/10.3390/jof8010029 - 29 Dec 2021

Cited by 1 | Viewed by 1727

Abstract

Purine auxotrophy is an abundant trait among eukaryotic parasites and a typical marker for many budding yeast strains. Supplementation with an additional purine source (such as adenine) is necessary to cultivate these strains. If not supplied in adequate amounts, purine starvation sets in. We explored purine starvation effects in a model organism, a budding yeast Saccharomyces cerevisiae ade8 knockout, at the level of cellular morphology, central carbon metabolism, and global transcriptome. We observed that purine-starved cells stopped their cycle in G1/G0 state and accumulated trehalose, and the intracellular concentration of AXP decreased, but adenylate charge remained stable. Cells became tolerant to severe environmental stresses. Intracellular RNA concentration decreased, and massive downregulation of ribosomal biosynthesis genes occurred. We proved that the expression of new proteins during purine starvation is critical for cells to attain stress tolerance phenotype Msn2/4p targets are upregulated in purine-starved cells when compared to cells cultivated in purine-rich media. The overall transcriptomic response to purine starvation resembles that of stationary phase cells. Our results demonstrate that the induction of a strong stress resistance phenotype in budding yeast can be caused not only by natural starvation, but also starvation for metabolic intermediates, such as purines. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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10 pages, 1941 KiB

Open AccessArticle

New Promoters for Metabolic Engineering of Ashbya gossypii

by Gloria Muñoz-Fernández, Javier-Fernando Montero-Bullón, José Luis Revuelta and Alberto Jiménez

J. Fungi 2021, 7(11), 906; https://0-doi-org.brum.beds.ac.uk/10.3390/jof7110906 - 26 Oct 2021

Cited by 4 | Viewed by 1909

Abstract

Ashbya gossypii is a filamentous fungus that is currently exploited for the industrial production of riboflavin. In addition, metabolically engineered strains of A. gossypii have also been described as valuable biocatalysts for the production of different metabolites such as folic acid, nucleosides, and biolipids. Hence, bioproduction in A. gossypii relies on the availability of well-performing gene expression systems both for endogenous and heterologous genes. In this regard, the identification of novel promoters, which are critical elements for gene expression, decisively helps to expand the A. gossypii molecular toolbox. In this work, we present an adaptation of the Dual Luciferase Reporter (DLR) Assay for promoter analysis in A. gossypii using integrative cassettes. We demonstrate the efficiency of the analysis through the identification of 10 new promoters with different features, including carbon source-regulatable abilities, that will highly improve the gene expression platforms used in A. gossypii. Three novel strong promoters (P_CCW12, P_SED1, and P_TSA1) and seven medium/weak promoters (P_HSP26, P_AGL366C, P_TMA10, P_CWP1, P_AFR038W, P_PFS1, and P_CDA2) are presented. The functionality of the promoters was further evaluated both for the overexpression and for the underexpression of the A. gossypii MSN2 gene, which induced significant changes in the sporulation ability of the mutant strains. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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16 pages, 2591 KiB

Open AccessArticle

Sterol Composition Modulates the Response of Saccharomyces cerevisiae to Iron Deficiency

by Tania Jordá, Nicolas Rozès and Sergi Puig

J. Fungi 2021, 7(11), 901; https://0-doi-org.brum.beds.ac.uk/10.3390/jof7110901 - 25 Oct 2021

Cited by 9 | Viewed by 2320

Abstract

Iron is a vital micronutrient that functions as an essential cofactor in multiple biological processes, including oxygen transport, cellular respiration, and metabolic pathways, such as sterol biosynthesis. However, its low bioavailability at physiological pH frequently leads to nutritional iron deficiency. The yeast Saccharomyces cerevisiae is extensively used to study iron and lipid metabolisms, as well as in multiple biotechnological applications. Despite iron being indispensable for yeast ergosterol biosynthesis and growth, little is known about their interconnections. Here, we used lipid composition analyses to determine that changes in the pattern of sterols impair the response to iron deprivation of yeast cells. Yeast mutants defective in ergosterol biosynthesis display defects in the transcriptional activation of the iron-acquisition machinery and growth defects in iron-depleted conditions. The transcriptional activation function of the iron-sensing Aft1 factor is interrupted due to its mislocalization to the vacuole. These data uncover novel links between iron and sterol metabolisms that need to be considered when producing yeast-derived foods or when treating fungal infections with drugs that target the ergosterol biosynthesis pathway. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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10 pages, 2879 KiB

Open AccessArticle

Perturbations in the Heme and Siroheme Biosynthesis Pathways Causing Accumulation of Fluorescent Free Base Porphyrins and Auxotrophy in Ogataea Yeasts

by Azamat V. Karginov, Alexander I. Alexandrov, Vitaly V. Kushnirov and Michael O. Agaphonov

J. Fungi 2021, 7(10), 884; https://0-doi-org.brum.beds.ac.uk/10.3390/jof7100884 - 19 Oct 2021

Cited by 4 | Viewed by 1827

Abstract

The biosynthesis of cyclic tetrapyrrol chromophores such as heme, siroheme, and chlorophyll involves the formation of fluorescent porphyrin precursors or compounds, which become fluorescent after oxidation. To identify Ogataea polymorpha mutations affecting the final steps of heme or siroheme biosynthesis, we performed a search for clones with fluorescence characteristic of free base porphyrins. One of the obtained mutants was defective in the gene encoding a homologue of Saccharomyces cerevisiae Met8 responsible for the last two steps of siroheme synthesis. Same as the originally obtained mutation, the targeted inactivation of this gene in O. polymorpha and O. parapolymorpha led to increased porphyrin fluorescence and methionine auxotrophy. These features allow the easy isolation of Met8-defective mutants and can potentially be used to construct auxotrophic strains in various yeast species. Besides MET8, this approach also identified the HEM3 gene encoding porphobilinogen deaminase, whose increased dosage led to free base porphyrin accumulation. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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25 pages, 5632 KiB

Open AccessArticle

Genome Comparisons of the Fission Yeasts Reveal Ancient Collinear Loci Maintained by Natural Selection

by Lajos Acs-Szabo, Laszlo Attila Papp, Matthias Sipiczki and Ida Miklos

J. Fungi 2021, 7(10), 864; https://0-doi-org.brum.beds.ac.uk/10.3390/jof7100864 - 14 Oct 2021

Cited by 1 | Viewed by 1881

Abstract

Fission yeasts have a unique life history and exhibit distinct evolutionary patterns from other yeasts. Besides, the species demonstrate stable genome structures despite the relatively fast evolution of their genomic sequences. To reveal what could be the reason for that, comparative genomic analyses were carried out. Our results provided evidence that the structural and sequence evolution of the fission yeasts were correlated. Moreover, we revealed ancestral locally collinear blocks (aLCBs), which could have been inherited from their last common ancestor. These aLCBs proved to be the most conserved regions of the genomes as the aLCBs contain almost eight genes/blocks on average in the same orientation and order across the species. Gene order of the aLCBs is mainly fission-yeast-specific but supports the idea of filamentous ancestors. Nevertheless, the sequences and gene structures within the aLCBs are as mutable as any sequences in other parts of the genomes. Although genes of certain Gene Ontology (GO) categories tend to cluster at the aLCBs, those GO enrichments are not related to biological functions or high co-expression rates, they are, rather, determined by the density of essential genes and Rec12 cleavage sites. These data and our simulations indicated that aLCBs might not only be remnants of ancestral gene order but are also maintained by natural selection. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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10 pages, 1354 KiB

Open AccessArticle

Regulation of Copper Metabolism by Nitrogen Utilization in Saccharomyces cerevisiae

by Suzie Kang, Hyewon Seo, Min-Gyu Lee and Cheol-Won Yun

J. Fungi 2021, 7(9), 756; https://0-doi-org.brum.beds.ac.uk/10.3390/jof7090756 - 14 Sep 2021

Cited by 1 | Viewed by 2012

Abstract

To understand the relationship between carbon or nitrogen utilization and iron homeostasis, we performed an iron uptake assay with several deletion mutants with partial defects in carbon or nitrogen metabolism. Among them, some deletion mutants defective in carbon metabolism partially and the MEP2 deletion mutant showed lower iron uptake activity than the wild type. Mep2 is known as a high-affinity ammonia transporter in Saccharomyces cerevisiae. Interestingly, we found that nitrogen starvation resulted in lower iron uptake activity than that of wild-type cells without downregulation of the genes involved in the high-affinity iron uptake system FET3/FTR1. However, the gene expression of FRE1 and CTR1 was downregulated by nitrogen starvation. The protein level of Ctr1 was also decreased by nitrogen starvation, and addition of copper to the nitrogen starvation medium partially restored iron uptake activity. However, the expression of MAC1, which is a copper-responsive transcriptional activator, was not downregulated by nitrogen starvation at the transcriptional level but was highly downregulated at the translational level. Mac1 was downregulated dramatically under nitrogen starvation, and treatment with MG132, which is an inhibitor of proteasome-dependent protein degradation, partially attenuated the downregulation of Mac1. Taken together, these results suggest that nitrogen starvation downregulates the high-affinity iron uptake system by degrading Mac1 in a proteasome-dependent manner and eventually downregulates copper metabolism. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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16 pages, 2191 KiB

Open AccessArticle

Acute Exposure to Bisphenol A Causes Oxidative Stress Induction with Mitochondrial Origin in Saccharomyces cerevisiae Cells

by Ivana Ďurovcová, Eduard Goffa, Zuzana Šestáková, Dominika Mániková, Katarína Gaplovská-Kyselá, Miroslav Chovanec and Andrea Ševčovičová

J. Fungi 2021, 7(7), 543; https://0-doi-org.brum.beds.ac.uk/10.3390/jof7070543 - 07 Jul 2021

Cited by 6 | Viewed by 2848

Abstract

Bisphenol A (BPA) is a major component of the most commonly used plastic products, such as disposable plastics, Tetra Paks, cans, sport protective equipment, or medical devices. Due to the accumulation of excessive amounts of plastic waste and the subsequent release of BPA into the environment, BPA is classified as a pollutant that is undesirable in the environment. To date, the most interesting finding is the ability of BPA to act as an endocrine disrupting compound due to its binding to estrogen receptors (ERs), and adverse physiological effects on living organisms may result from this action. Since evidence of the potential pro-oxidizing effects of BPA has accumulated over the last years, herein, we focus on the detection of oxidative stress and its origin following BPA exposure using pulsed-field gel electrophoresis, flow cytometry, fluorescent microscopy, and Western blot analysis. Saccharomyces cerevisiae cells served as a model system, as these cells lack ERs allowing us to dissect the ER-dependent and -independent effects of BPA. Our data show that high concentrations of BPA affect cell survival and cause increased intracellular oxidation in yeast, which is primarily generated in the mitochondrion. However, an acute BPA exposure does not lead to significant oxidative damage to DNA or proteins. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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Review

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22 pages, 1299 KiB

Open AccessReview

Metabolic Engineering Strategies for Improved Lipid Production and Cellular Physiological Responses in Yeast Saccharomyces cerevisiae

by Wei Jiang, Chao Li, Yanjun Li and Huadong Peng

J. Fungi 2022, 8(5), 427; https://0-doi-org.brum.beds.ac.uk/10.3390/jof8050427 - 21 Apr 2022

Cited by 9 | Viewed by 3854

Abstract

Microbial lipids have been a hot topic in the field of metabolic engineering and synthetic biology due to their increased market and important applications in biofuels, oleochemicals, cosmetics, etc. This review first compares the popular hosts for lipid production and explains the four modules for lipid synthesis in yeast, including the fatty acid biosynthesis module, lipid accumulation module, lipid sequestration module, and fatty acid modification module. This is followed by a summary of metabolic engineering strategies that could be used for enhancing each module for lipid production. In addition, the efforts being invested in improving the production of value-added fatty acids in engineered yeast, such as cyclopropane fatty acid, ricinoleic acid, gamma linoleic acid, EPA, and DHA, are included. A discussion is further made on the potential relationships between lipid pathway engineering and consequential changes in cellular physiological properties, such as cell membrane integrity, intracellular reactive oxygen species level, and mitochondrial membrane potential. Finally, with the rapid development of synthetic biology tools, such as CRISPR genome editing tools and machine learning models, this review proposes some future trends that could be employed to engineer yeast with enhanced intracellular lipid production while not compromising much of its cellular health. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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18 pages, 1947 KiB

Open AccessReview

Differential Interactions of Molecular Chaperones and Yeast Prions

by Yury A. Barbitoff, Andrew G. Matveenko and Galina A. Zhouravleva

J. Fungi 2022, 8(2), 122; https://0-doi-org.brum.beds.ac.uk/10.3390/jof8020122 - 27 Jan 2022

Cited by 5 | Viewed by 2661

Abstract

Baker’s yeast Saccharomyces cerevisiae is an important model organism that is applied to study various aspects of eukaryotic cell biology. Prions in yeast are self-perpetuating heritable protein aggregates that can be leveraged to study the interaction between the protein quality control (PQC) machinery and misfolded proteins. More than ten prions have been identified in yeast, of which the most studied ones include [PSI⁺], [URE3], and [PIN⁺]. While all of the major molecular chaperones have been implicated in propagation of yeast prions, many of these chaperones differentially impact propagation of different prions and/or prion variants. In this review, we summarize the current understanding of the life cycle of yeast prions and systematically review the effects of different chaperone proteins on their propagation. Our analysis clearly shows that Hsp40 proteins play a central role in prion propagation by determining the fate of prion seeds and other amyloids. Moreover, direct prion-chaperone interaction seems to be critically important for proper recruitment of all PQC components to the aggregate. Recent results also suggest that the cell asymmetry apparatus, cytoskeleton, and cell signaling all contribute to the complex network of prion interaction with the yeast cell. Full article

(This article belongs to the Special Issue Yeast Genetics 2021)

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