scholarly journals Himalayan Saccharomyces eubayanus Genome Sequences Reveal Genetic Markers Explaining Heterotic Maltotriose Consumption by Saccharomyces pastorianus Hybrids

2019 ◽  
Vol 85 (22) ◽  
Author(s):  
Nick Brouwers ◽  
Anja Brickwedde ◽  
Arthur R. Gorter de Vries ◽  
Marcel van den Broek ◽  
Susan M. Weening ◽  
...  

ABSTRACT Saccharomyces pastorianus strains are hybrids of Saccharomyces cerevisiae and Saccharomyces eubayanus that have been domesticated for centuries in lager beer brewing environments. As sequences and structures of S. pastorianus genomes are being resolved, molecular mechanisms and evolutionary origins of several industrially relevant phenotypes remain unknown. This study investigates how maltotriose metabolism, a key feature in brewing, may have arisen in early S. eubayanus × S. cerevisiae hybrids. To address this question, we generated a nearly complete genome assembly of Himalayan S. eubayanus strains of the Holarctic subclade. This group of strains has been proposed to be the S. eubayanus subgenome origin of current S. pastorianus strains. The Himalayan S. eubayanus genomes harbored several copies of an S. eubayanus AGT1 (SeAGT1) α-oligoglucoside transporter gene with high sequence identity to genes encountered in S. pastorianus. Although Himalayan S. eubayanus strains cannot grow on maltose and maltotriose, their maltose-hydrolase and SeMALT1 and SeAGT1 maltose transporter genes complemented the corresponding null mutants of S. cerevisiae. Expression, in Himalayan S. eubayanus of a functional S. cerevisiae maltose metabolism regulator gene (MALx3) enabled growth on oligoglucosides. The hypothesis that the maltotriose-positive phenotype in S. pastorianus is a result of heterosis was experimentally tested by constructing an S. cerevisiae × S. eubayanus laboratory hybrid with a complement of maltose metabolism genes that resembles that of current S. pastorianus strains. The ability of this hybrid to consume maltotriose in brewer’s wort demonstrated regulatory cross talk between subgenomes and thereby validated this hypothesis. These results support experimentally the new postulated hypothesis on the evolutionary origin of an essential phenotype of lager brewing strains and valuable knowledge for industrial exploitation of laboratory-made S. pastorianus-like hybrids. IMPORTANCE S. pastorianus, an S. cerevisiae × S. eubayanus hybrid, is used for production of lager beer, the most produced alcoholic beverage worldwide. It emerged by spontaneous hybridization and colonized early lager brewing processes. Despite accumulation and analysis of genome sequencing data of S. pastorianus parental genomes, the genetic blueprint of industrially relevant phenotypes remains unresolved. Assimilation of maltotriose, an abundant sugar in wort, has been postulated to be inherited from the S. cerevisiae parent. Here, we demonstrate that although Asian S. eubayanus isolates harbor a functional maltotriose transporter SeAGT1 gene, they are unable to grow on α-oligoglucosides, but expression of S. cerevisiae regulator MAL13 (ScMAL13) was sufficient to restore growth on trisaccharides. We hypothesized that the S. pastorianus maltotriose phenotype results from regulatory interaction between S. cerevisiae maltose transcription activator and the promoter of SeAGT1. We experimentally confirmed the heterotic nature of the phenotype, and thus these results provide experimental evidence of the evolutionary origin of an essential phenotype of lager brewing strains.

2019 ◽  
Author(s):  
Nick Brouwers ◽  
Anja Brickwedde ◽  
Arthur R. Gorter de Vries ◽  
Marcel van den Broek ◽  
Susan M. Weening ◽  
...  

AbstractS. pastorianus strains are hybrids of S. cerevisiae and S. eubayanus that have been domesticated for several centuries in lager-beer brewing environments. As sequences and structures of S. pastorianus genomes are being resolved, molecular mechanisms and evolutionary origin of several industrially relevant phenotypes remain unknown. This study investigates how maltotriose metabolism, a key feature in brewing, may have arisen in early S. eubayanus × S. cerevisiae hybrids. To address this question, we generated a near-complete genome assembly of Himalayan S. eubayanus strains of the Holarctic subclade. This group of strains have been proposed to be the origin of the S. eubayanus subgenome of current S. pastorianus strains. The Himalayan S. eubayanus genomes harbored several copies of a SeAGT1 α-oligoglucoside transporter gene with high sequence identity to genes encountered in S. pastorianus. Although Himalayan S. eubayanus strains are unable to grown on maltose and maltotriose, their maltose-hydrolase and SeMALT1 and SeAGT1 maltose-transporter genes complemented the corresponding null mutants of S. cerevisiae. Expression, in a Himalayan S. eubayanus strain, of a functional S. cerevisiae maltose-metabolism regulator gene (MALx3) enabled growth on oligoglucosides. The hypothesis that the maltotriose-positive phenotype in S. pastorianus is a result of heterosis was experimentally tested by constructing a S. cerevisiae × S. eubayanus laboratory hybrid with a complement of maltose-metabolism genes that resembles that of current S. pastorianus strains. The ability of this hybrid to consume maltotriose in brewer’s wort demonstrated regulatory cross talk between sub-genomes and thereby validated this hypothesis. These results provide experimental evidence of the evolutionary origin of an essential phenotype of lager-brewing strains and valuable knowledge for industrial exploitation of laboratory-made S. pastorianus-like hybrids.ImportanceS.pastorianus, a S.cerevisiae X S.eubayanus hybrid, is used for production of lager beer, the most produced alcoholic beverage worldwide It emerged by spontaneous hybridization and have colonized early lager-brewing processes. Despite accumulation and analysis of genome sequencing data of S.pastorianus parental genomes, the genetic blueprint of industrially relevant phenotypes remains unknown. Assimilation of wort abundant sugar maltotriose has been postulated to be inherited from S.cerevisiae parent. Here, we demonstrate that although Asian S.eubayanus isolates harbor a functional maltotriose transporter SeAGT1 gene, they are unable to grow on α-oligoglucosides, but expression of S. cerevisae regulator ScMAL13 was sufficient to restore growth on trisaccharides. We hypothesized that S. pastorianus maltotriose phenotype results from regulatory interaction between S.cerevisae maltose transcription activator and the promoter of SeAGT1. We experimentally confirmed the heterotic nature of the phenotype and thus this results provide experimental evidence of the evolutionary origin of an essential phenotype of lager-brewing strains.


2015 ◽  
Vol 81 (23) ◽  
pp. 8202-8214 ◽  
Author(s):  
Stijn Mertens ◽  
Jan Steensels ◽  
Veerle Saels ◽  
Gert De Rouck ◽  
Guido Aerts ◽  
...  

ABSTRACTLager beer is the most consumed alcoholic beverage in the world. Its production process is marked by a fermentation conducted at low (8 to 15°C) temperatures and by the use ofSaccharomyces pastorianus, an interspecific hybrid betweenSaccharomyces cerevisiaeand the cold-tolerantSaccharomyces eubayanus. Recent whole-genome-sequencing efforts revealed that the currently available lager yeasts belong to one of only two archetypes, “Saaz” and “Frohberg.” This limited genetic variation likely reflects that all lager yeasts descend from only two separate interspecific hybridization events, which may also explain the relatively limited aromatic diversity between the available lager beer yeasts compared to, for example, wine and ale beer yeasts. In this study, 31 novel interspecific yeast hybrids were developed, resulting from large-scale robot-assisted selection and breeding between carefully selected strains ofS. cerevisiae(six strains) andS. eubayanus(two strains). Interestingly, many of the resulting hybrids showed a broader temperature tolerance than their parental strains and referenceS. pastorianusyeasts. Moreover, they combined a high fermentation capacity with a desirable aroma profile in laboratory-scale lager beer fermentations, thereby successfully enriching the currently available lager yeast biodiversity. Pilot-scale trials further confirmed the industrial potential of these hybrids and identified one strain, hybrid H29, which combines a fast fermentation, high attenuation, and the production of a complex, desirable fruity aroma.


2017 ◽  
Vol 199 (16) ◽  
Author(s):  
Yiwei Meng ◽  
Zhou Yang ◽  
Bin Cheng ◽  
Xinyu Nie ◽  
Shannan Li ◽  
...  

ABSTRACT Two NhaD-type antiporters, NhaD1 and NhaD2, from the halotolerant and alkaliphilic Halomonas sp. strain Y2, exhibit different physiological functions in regard to Na+ and Li+ resistance, although they share high sequence identity. In the present study, the truncation of an additional 4 C-terminal residues from NhaD2 or an exchange of 39 N-terminal residues between these proteins resulted in the complete loss of antiporter activity. Interestingly, combining 39 N-terminal residues and 7 C-terminal residues of NhaD2 (N39D2-C7) partially recovered the activity for Na+ and Li+ expulsion, as well as complementary growth following exposure to 300 mM Na+ and 100 mM Li+ stress. The recovered activity of chimera N39D2-C7 indicated that the N and C termini are structurally dependent on each other and function synergistically. Furthermore, fluorescence resonance energy transfer (FRET) analysis suggested that the N and C termini are relatively close in proximity which may account for their synergistic function in ion translocation. In the N-terminal region of N39D2-C7, the replacement of Glu38 with Pro abolished the recovered complementary and transport activities. In addition, this amino acid substitution in NhaD2 resulted in a drastically decreased complementation ability in Escherichia coli KNabc (level identical to that of NhaD1), as well as decreased activity and an altered pH profile. IMPORTANCE Limited information on NhaD antiporters supports speculation that these antiporters are important for resistance to high salinity and alkalinity. Moreover, only a few functional residues have been identified in NhaD antiporters, and there is limited literature on the molecular mechanisms of NhaD antiporter activity. The altered antiporter abilities of chimeras and mutants in this study implicate the functions of the N and C termini, especially Glu38, in pH regulation and ion translocation, and, most importantly, the essential roles of this negatively charged residue in maintaining the physiological function of NhaD2. These findings further our understanding of the molecular mechanism of NhaD antiporters for ion transport.


2014 ◽  
Vol 13 (10) ◽  
pp. 1256-1265 ◽  
Author(s):  
Jürgen Wendland

ABSTRACTAlcoholic fermentations have accompanied human civilizations throughout our history. Lager yeasts have a several-century-long tradition of providing fresh beer with clean taste. The yeast strains used for lager beer fermentation have long been recognized as hybrids between twoSaccharomycesspecies. We summarize the initial findings on this hybrid nature, the genomics/transcriptomics of lager yeasts, and established targets of strain improvements. Next-generation sequencing has provided fast access to yeast genomes. Its use in population genomics has uncovered many more hybridization events withinSaccharomycesspecies, so that lager yeast hybrids are no longer the exception from the rule. These findings have led us to propose network evolution withinSaccharomycesspecies. This “web of life” recognizes the ability of closely related species to exchange DNA and thus drain from a combined gene pool rather than be limited to a gene pool restricted by speciation. Within the domesticated lager yeasts, two groups, the Saaz and Frohberg groups, can be distinguished based on fermentation characteristics. Recent evidence suggests that these groups share an evolutionary history. We thus propose to refer to the Saaz group asSaccharomyces carlsbergensisand to the Frohberg group asSaccharomyces pastorianusbased on their distinct genomes. New insight into the hybrid nature of lager yeast will provide novel directions for future strain improvement.


Processes ◽  
2021 ◽  
Vol 9 (9) ◽  
pp. 1628
Author(s):  
Massimo Iorizzo ◽  
Francesco Letizia ◽  
Gianluca Albanese ◽  
Francesca Coppola ◽  
Angelita Gambuti ◽  
...  

Saccharomyces pastorianus, genetic hybrids of Saccharomyces cerevisiae and the Saccharomyces eubayanus, is one of the most widely used lager yeasts in the brewing industry. In recent years, new strategies have been adopted and new lines of research have been outlined to create and expand the pool of lager brewing starters. The vineyard microbiome has received significant attention in the past few years due to many opportunities in terms of biotechnological applications in the winemaking processes. However, the characterization of S. cerevisiae strains isolated from winery environments as an approach to selecting starters for beer production has not been fully investigated, and little is currently available. Four wild cryotolerant S. cerevisiae strains isolated from vineyard environments were evaluated as potential starters for lager beer production at laboratory scale using a model beer wort (MBW). In all tests, the industrial lager brewing S. pastorianus Weihenstephan 34/70 was used as a reference strain. The results obtained, although preliminary, showed some good properties of these strains, such as antioxidant activity, flocculation capacity, efficient fermentation at 15 °C and low diacetyl production. Further studies will be carried out using these S. cerevisiae strains as starters for lager beer production on a pilot scale in order to verify the chemical and sensory characteristics of the beers produced.


2019 ◽  
Vol 5 (1) ◽  
pp. eaav1869 ◽  
Author(s):  
EmilyClare P. Baker ◽  
David Peris ◽  
Ryan V. Moriarty ◽  
Xueying C. Li ◽  
Justin C. Fay ◽  
...  

A growing body of research suggests that the mitochondrial genome (mtDNA) is important for temperature adaptation. In the yeast genusSaccharomyces, species have diverged in temperature tolerance, driving their use in high- or low-temperature fermentations. Here, we experimentally test the role of mtDNA in temperature tolerance in synthetic and industrial hybrids (Saccharomyces cerevisiae×Saccharomyces eubayanusorSaccharomyces pastorianus), which cold-brew lager beer. We find that the relative temperature tolerances of hybrids correspond to the parent donating mtDNA, allowing us to modulate lager strain temperature preferences. The strong influence of mitotype on the temperature tolerance of otherwise identical hybrid strains provides support for the mitochondrial climactic adaptation hypothesis in yeasts and demonstrates how mitotype has influenced the world’s most commonly fermented beverage.


2014 ◽  
Vol 82 (5) ◽  
pp. 1744-1754 ◽  
Author(s):  
Tram N. Cao ◽  
Zhuyun Liu ◽  
Tran H. Cao ◽  
Kathryn J. Pflughoeft ◽  
Jeanette Treviño ◽  
...  

ABSTRACTDespite the public health challenges associated with the emergence of new pathogenic bacterial strains and/or serotypes, there is a dearth of information regarding the molecular mechanisms that drive this variation. Here, we began to address the mechanisms behind serotype-specific variation between serotype M1 and M3 strains of the human pathogenStreptococcus pyogenes(the group AStreptococcus[GAS]). Spatially diverse contemporary clinical serotype M3 isolates were discovered to contain identical inactivating mutations within genes encoding two regulatory systems that control the expression of important virulence factors, including the thrombolytic agent streptokinase, the protease inhibitor-binding protein-G-related α2-macroglobulin-binding (GRAB) protein, and the antiphagocytic hyaluronic acid capsule. Subsequent analysis of a larger collection of isolates determined that M3 GAS, since at least the 1920s, has harbored a 4-bp deletion in thefasCgene of thefasBCAXregulatory system and an inactivating polymorphism in therivRregulator-encoding gene. ThefasCandrivRmutations in M3 isolates directly affect the virulence factor profile of M3 GAS, as evident by a reduction in streptokinase expression and an enhancement of GRAB expression. Complementation of thefasCmutation in M3 GAS significantly enhanced levels of the small regulatory RNA FasX, which in turn enhanced streptokinase expression. Complementation of therivRmutation in M3 GAS restored the regulation ofgrabmRNA abundance but did not alter capsule mRNA levels. While important, thefasCandrivRmutations do not provide a full explanation for why serotype M3 strains are associated with unusually severe invasive infections; thus, further investigation is warranted.


2016 ◽  
Vol 84 (6) ◽  
pp. 1879-1886 ◽  
Author(s):  
Lena J. Heung ◽  
Tobias M. Hohl

Cryptococcus neoformansis an opportunistic fungal pathogen that is inhaled into the lungs and can lead to life-threatening meningoencephalitis in immunocompromised patients. Currently, the molecular mechanisms that regulate the mammalian immune response to respiratory cryptococcal challenge remain poorly defined. DAP12, a signaling adapter for multiple pattern recognition receptors in myeloid and natural killer (NK) cells, has been shown to play both activating and inhibitory roles during lung infections by different bacteria and fungi. In this study, we demonstrate that DAP12 plays an important inhibitory role in the immune response toC. neoformans. Infectious outcomes in DAP12−/−mice, including survival and lung fungal burden, are significantly improved compared to those in C57BL/6 wild-type (WT) mice. We find that eosinophils and macrophages are decreased while NK cells are increased in the lungs of infected DAP12−/−mice. In contrast to WT NK cells, DAP12−/−NK cells are able to repressC. neoformansgrowthin vitro. Additionally, DAP12−/−macrophages are more highly activated than WT macrophages, with increased production of tumor necrosis factor (TNF) and CCL5/RANTES and more efficient uptake and killing ofC. neoformans. These findings suggest that DAP12 acts as a brake on the pulmonary immune response toC. neoformansby promoting pulmonary eosinophilia and by inhibiting the activation and antifungal activities of effector cells, including NK cells and macrophages.


2021 ◽  
Vol ahead-of-print (ahead-of-print) ◽  
Author(s):  
Simone Sehnem ◽  
Ana Beatriz Lopes de Sousa Jabbour ◽  
Diogo Amarildo da Conceição ◽  
Darciana Weber ◽  
Dulcimar José Julkovski

PurposeThere is no literature that sought the revision and integration of ecological modernization of theory (EMT) and circular economy (CE). This article aims to answer the questions: How have companies developed circular economy practices within their operations? Why have companies invested in circular economy practices? Based on the answers to these questions, it would be possible to understand how the assumptions of EMT explain the adoption of CE practices.Design/methodology/approachThe research method is based on multiple case studies with Brazilian craft brewery companies.FindingsCompanies studied have pursed the adoption of circular economy practices, but the full potential of the use of biological cycles within their processes of production has been hampered by the lack of external policies and clear governance toward circular economy. Thus, pollution prevention policies may be an important driver for organizations be able to advance in circular economy practices.Research limitations/implicationsResearch limitations are coverage only of organizations located in an emerging country. A second limitation is that the data saturation was partially achieved in some of the analyzed cases, especially by the limitation of the time period surveyed (transversal research). The longitudinal evaluation is timely for circular economy surveys.Practical implicationsThe formulation of a regulatory framework through a participative, interactive and decentralized process, capable of delegating responsibilities and incentives to all entities, to contemplate a tax restructuring for the alcoholic beverage sector, could corroborate to create a new standard of action for the sector.Social implicationsThis document highlights how the principles of ecological modernization can contribute to the advancement of the circular economy. The understanding can help in the design of processes that provide circular and sustainable operations as a social good in a effective value.Originality/valueThe principles from EMT, such as the development of an institutional environment towards corporate environmental proactive, could enhance the adoption of circular economy practices within the brewery sector. Despite of the class association of the brewery sector has supported the companies studied to move forward in other matters, a clear governance is necessary to guide organizations towards circular economy practices.


2018 ◽  
Vol 85 (10) ◽  
Author(s):  
Alli Lynch ◽  
Seshu R. Tammireddy ◽  
Mary K. Doherty ◽  
Phillip D. Whitfield ◽  
David J. Clarke

ABSTRACTAcylated amino acids function as important components of the cellular membrane in some bacteria. Biosynthesis is initiated by theN-acylation of the amino acid, and this is followed by subsequentO-acylation of the acylated molecule, resulting in the production of the mature diacylated amino acid lipid. In this study, we use both genetics and liquid chromatography-mass spectrometry (LC-MS) to characterize the biosynthesis and function of a diacylated glycine lipid (GL) species produced inBacteroides thetaiotaomicron. We, and others, have previously reported the identification of a gene, namedglsBin this study, that encodes anN-acyltransferase activity responsible for the production of a monoacylated glycine calledN-acyl-3-hydroxy-palmitoyl glycine (or commendamide). In all of theBacteroidalesgenomes sequenced so far, theglsBgene is located immediately downstream from a gene, namedglsA, that is also predicted to encode a protein with acyltransferase activity. We use LC-MS to show that the coexpression ofglsBandglsAresults in the production of GL inEscherichia coli. We constructed a deletion mutant of theglsBgene inB. thetaiotaomicron, and we confirm thatglsBis required for the production of GL inB. thetaiotaomicron. Moreover, we show thatglsBis important for the ability ofB. thetaiotaomicronto adapt to stress and colonize the mammalian gut. Therefore, this report describes the genetic requirements for the biosynthesis of GL, a diacylated amino acid species that contributes to fitness in the human gut bacteriumB. thetaiotaomicron.IMPORTANCEThe gut microbiome has an important role in both health and disease of the host. The mammalian gut microbiome is often dominated by bacteria from theBacteroidales, an order that includesBacteroidesandPrevotella. In this study, we have identified an acylated amino acid, called glycine lipid, produced byBacteroides thetaiotaomicron, a beneficial bacterium originally isolated from the human gut. In addition to identifying the genes required for the production of glycine lipids, we show that glycine lipids have an important role during the adaptation ofB. thetaiotaomicronto a number of environmental stresses, including exposure to either bile or air. We also show that glycine lipids are important for the normal colonization of the murine gut byB. thetaiotaomicron. This work identifies glycine lipids as an important fitness determinant inB. thetaiotaomicronand therefore increases our understanding of the molecular mechanisms underpinning colonization of the mammalian gut by beneficial bacteria.


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