Biology subjects

Villarreal, P.

Publications and source records attributed to Villarreal, P..

9 recordsLinked to original sources

bioRxiv2026

Convergent targeting of conserved regulatory networks during thermal evolution across Saccharomyces

Whether evolution follows predictable genetic paths across species remains a central question in evolutionary biology, particularly as rising temperatures reshape species distributions worldwide. Despite its importance, the genetic basis of thermal adaptation remains poorly understood across divergent species. Here, we use the yeast genus Saccharomyces as a comparative model to investigate how species with contrasting thermal niches adapt to rising temperatures. We combined experimental evolution under progressively increasing temperatures for up to [~]600 generations with whole-genome sequencing of 256 evolved genotypes, followed by transcriptomic, functional, and physiological analyses across eight species. Despite large differences in ancestral thermal tolerance and evolutionary outcomes, selection repeatedly targeted the same conserved regulatory pathways across species. Independent lineages accumulated de novo mutations in central growth and stress response networks, particularly in TORC1, PKA, and MAPK signaling pathways, revealing striking molecular convergence across species occupying distinct thermal environments. However, these shared genetic targets produced divergent transcriptional and physiological responses depending on species background, indicating that thermal adaptation primarily proceeds through rewiring of conserved regulatory hubs rather than changes in temperature-specific enzymes. Cold-tolerant species frequently lost mitochondrial DNA during thermal evolution, yet loss alone was insufficient to reproduce the adaptive thermal phenotypes of evolved populations. Together, our results show that adaptation to increasing temperature is driven by predictable changes in conserved regulatory networks, while species-specific constraints shape divergent phenotypic outcomes. These findings reveal both the predictability and contingency of evolutionary responses to rising temperature across species.

genomics↗

bioRxiv2026

Host-shift adaptation shapes genome architecture in S. eubayanus

The host environment can profoundly shape the genome architecture of microbial species, and Saccharomyces eubayanus, the wild progenitor of lager yeast, provides a natural system to study this process. Most populations are associated with Nothofagus trees across Patagonia, whereas related Holarctic strains occur in the Northern Hemisphere and are associated with non-Nothofagus hosts. The evolutionary events leading to the emergence of these northern populations on a novel host remain unclear. Here, we analyzed 471 genomes from eight countries and different hosts, Nothofagus in the Southern Hemisphere and non-Nothofagus tree species in the Northern Hemisphere. Phylogenomic analysis identified eight Patagonian lineages and revealed that Holarctic strains derived from recent admixture among Patagonian ancestors, generating the genomic background of the lager-yeast mother lineage. Long-read assemblies showed that non-Nothofagus-associated strains harbor an elevated burden of structural variants (SVs), particularly in subtelomeric MAL and IMA regions, involved in sugar metabolism. Phenotypic tests confirmed that Nothofagus isolates efficiently metabolize maltose, while non-Nothofagus strains do not, a pattern linked to recurrent SVs and loss-of-function mutations in MAL33. Consistently, bark-sugar profiling revealed that maltose is abundant in Nothofagus but absent in non-Nothofagus hosts, providing an ecological context for these genomic and phenotypic differences. These results support a model in which northward dispersal of Patagonian lineages into non-Nothofagus forests enriched admixed genotypes, generating genomic mosaics that accumulated structural changes and losses in maltose utilization. This interplay between gene flow and genome flexibility enabled host switching and global expansion, illustrating how ecological transitions reorder genomes and drive microbial diversification.

evolutionary biology↗

bioRxiv2025

Contrasting evolutionary forces of specialization and admixture underlie the genomic and phenotypic diversity of Yarrowia lipolytica

Microbial diversity emerges from evolutionary processes that shape genomic and phenotypic traits in response to complex environmental pressures. Deciphering these dynamics is key to understanding microbial ecology and advancing biotechnological applications. Here, we use the yeast Yarrowia lipolytica to study genomic signatures of adaptation across a broad range of environments and to illustrate how population-level data can inform targeted bioprospecting for industrial traits. Whole-genome and phenotypic analyses of 126 isolates from natural and anthropogenic environments reveal a complex population structure in this species, shaped by both niche specialization and admixture events. Structured lineages exhibit ecological filtering, reduced genetic diversity, and distinct gene content, consistent with adaptation to substrates like dairy, hydrocarbons, or industrial substrates. In contrast, admixed populations display greater genetic diversity and broader phenotypic capacity, including enhanced stress tolerance and metabolic flexibility. Genome plasticity, reflected in pangenome and CNV variation, aligns with ecological origin, while trait assays link phenotypic divergence to underlying genetic variation. For instance, better growth performance on acetate, an ecological and industrially relevant trait, is associated with hydrocarbon-adapted strains and likely linked to variation in regulatory regions of acetate metabolism genes. Together, our results reflect how divergent evolutionary trajectories--ranging from ecological specialization to genomic plasticity through admixture-- underpin the species ecological success and provide a framework for harnessing its natural diversity in microbial bioprospecting.

evolutionary biology↗

bioRxiv2025

Transcriptional Reprogramming Drives Cold Adaptation During Long-Term Starvation in S. eubayanus

The ability of microorganisms to survive prolonged periods of nutrient scarcity is essential for their survival. Yet, the underlying adaptive mechanisms remain partly understood, especially in non-model eukaryotes. Here, we examined how the cryotolerant yeast Saccharomyces eubayanus adapts to 60 days of cold (4{degrees}C) starvation, focusing on the roles of genetic and transcriptional changes. We find that the primary engine of the long-term adaptation is a stable, reprogrammed transcriptional state, rather than the selection of point mutations. Aged isolates exhibited improved growth performance and cryotolerance, a phenotype that remained stable for [~]40 generations. This specialist adaptation involved a trade-off with tolerance to other stresses. Whole-genome sequencing revealed few fixed mutations, indicating that genetic variation did not drive the phenotype. Transcriptomic analysis revealed a significant physiological reprogramming, with cells shifting from anabolic activity to a catabolic, scavenging state driven by enhanced respiration and activation of the General Stress Response. This work highlights that a stable transcriptional state drives long-term cold adaptation, providing the foundation for the superior phenotype of aged isolates that persist through generations.

microbiology↗

bioRxiv2025

Allelic variation in MAL33 drives ecological adaptation of maltose metabolism in Saccharomyces eubayanus

Maltose is one of the most abundant sugars in brewers wort, and its efficient utilization is critical for successful fermentation. However, maltose consumption varies naturally among Saccharomyces eubayanus strains isolated from different host trees, such as Quercus and Nothofagus. To identify the genetic determinants underlying these phenotypic differences, we performed bulk segregant analysis (BSA) and quantitative trait loci (QTL) mapping using an F2 offspring derived from QC18 (Quercus-associated) and CL467.1 (Nothofagus-associated) strains. QTL mapping identified two significant genomic regions on subtelomeric loci of chromosomes V-R and XVI-L, each containing complete MAL loci composed of MAL32 (encoding maltase), MAL31 (transporter), and MAL33 (transcriptional activator) genes. Comparative polymorphism analyses identified mutations in MAL32 and MAL33 of QC18, including frameshift mutations resulting in premature stop codons. Functional validation demonstrated that the heterologous expression of MAL33ChrV from CL467.1 fully restored maltose utilization in QC18, indicating the functional presence of MAL33 cis-regulatory sequences and MAL32 and MAL31 genes in QC18. While structural protein predictions identified truncation and impaired functionality in the maltose-responsive activation domain of Mal33p from QC18, overexpression of QC18s own MAL33ChrV allele also improved maltose metabolism, suggesting dosage-dependent transcriptional limitations rather than complete functional loss. These results indicate that allelic variations in the maltose-responsive activation domain of Mal33p lead to differences in maltose consumption between strains. We hypothesized that reduced maltose metabolism in QC18 is an adaptive response to the distinct sugar composition in Quercus robur bark, contrasting with the starch-rich environment of Nothofagus pumilio. These findings highlight subtelomeric MAL gene diversity as a reservoir of evolutionary plasticity, representing a key evolutionary mechanism that influences maltose adaptation among natural Saccharomyces isolates.

evolutionary biology↗

bioRxiv2024

An integrative taxonomy approach reveals Saccharomyces chiloensis sp. nov. as a newly discovered species from Coastal Patagonia

Species delineation in microorganisms is challenging due to the limited markers available for accurate species assignment. Here, we applied an integrative taxonomy approach, combining extensive sampling, whole-genome sequence-based classification, phenotypic profiling, and assessment of interspecific reproductive isolation. Our work reveals the presence of a distinct Saccharomyces lineage in Nothofagus forests of coastal Patagonia. This lineage, designated Saccharomyces chiloensis sp. nov., exhibits 7% genetic divergence from its sister species S. uvarum, as revealed by whole-genome sequencing and population analyses. The South America-C (SA-C) coastal Patagonia population forms a unique clade closely related to a previously described divergent S. uvarum population from Oceania (AUS, found in Australia and New Zealand). Our species reclassification is supported by a low Ortho Average Nucleotide Identity (OANI) of 93% in SA-C and AUS relative to S. uvarum, which falls below the suggested species delineation threshold of 95%, indicating an independent evolutionary lineage. Hybrid spore viability assessment provided compelling evidence that SA-C and AUS are reproductively isolated from S. uvarum. In addition, we found unique structural variants between S. chiloensis sp. nov. lineages, including large-scale chromosomal translocations and inversions, together with a distinct phenotypic profile, emphasizing their intraspecies genetic distinctiveness. We suggest that S. chiloensis sp. nov diverged from S. uvarum in allopatry due to glaciation, followed by post-glacial dispersal, resulting in distinct lineages on opposite sides of the Pacific Ocean. The discovery of S. chiloensis sp. nov. illustrates the uniqueness of Patagonias coastal biodiversity and underscores the importance of adopting an integrative taxonomic approach in species delineation to unveil cryptic microbial species. The holotype of S. chiloensis sp. nov. is CBS 18620T.

genetics↗

bioRxiv2024

Wild Patagonian yeast improve the evolutionary potential of novel interspecific hybrid strains for Lager brewing

ABSTRACTLager yeasts are limited to a few strains worldwide, imposing restrictions on flavour and aroma diversity and hindering our understanding of the complex evolutionary mechanisms during yeast domestication. The recent finding of diverse S. eubayanus lineages from Patagonia offers potential for generating new Lager yeasts and obtaining insights into the domestication process. Here, we leverage the natural genetic diversity of S. eubayanus and expand the Lager yeast repertoire by including three distinct Patagonian S. eubayanus lineages. We used experimental evolution and selection on desirable traits to enhance the fermentation profiles of novel S. cerevisiae x S. eubayanus hybrids. Our analyses reveal an intricate interplay of pre-existing diversity, selection on species-specific mitochondria, de-novo mutations, and gene copy variations in sugar metabolism genes, resulting in high ethanol production and unique aroma profiles. Hybrids with S. eubayanus mitochondria exhibited greater evolutionary potential and superior fitness post-evolution, analogous to commercial Lager hybrids. Using genome-wide screens of the parental subgenomes, we identified genetic changes in IRA2, SNF3, IMA1 and MALX genes that influence maltose metabolism, and increase glycolytic flux and sugar consumption in the evolved hybrids. Functional validation and transcriptome analyses confirmed increased maltose-related gene expression, influencing grater maltotriose consumption in evolved hybrids. This study demonstrates the potential for generating industrially viable Lager yeast hybrids from wild Patagonian strains. Our hybridization, evolution, and mitochondrial selection approach produced hybrids with high fermentation capacity, expands Lager beer brewing options, and deepens our knowledge of Lager yeast domestication.

microbiology↗

bioRxiv2023

Domestication signatures in the non-conventional yeast Lachancea cidri

Evaluating domestication signatures beyond model organisms is essential for thoroughly understanding the genotype-phenotype relationship in wild and human-related environments. Structural variations (SVs) can significantly impact phenotypes playing an important role in the physiological adaptation of species to different niches, including during domestication. A detailed characterization of the fitness consequences of these genomic rearrangements, however, is still limited in non-model systems, largely due to the paucity of direct comparisons between domesticated and wild isolates. Here, we used a combination of sequencing strategies to explore major genomic rearrangements in a Lachancea cidri yeast strain isolated from cider (CBS2950) and compared them to those in eight wild isolates from primary forests. Genomic analysis revealed dozens of SVs, including a large reciprocal translocation ([~]16 kb and 500 kb) present in the cider strain, but absent from all wild strains. Interestingly, the number of SVs was higher relative to single-nucleotide polymorphisms in the cider strain, suggesting a significant role on the strains phenotypic variation. The set of SVs identified directly impacts dozens of genes, and likely underpins the greater fermentation performance in the L. cidri CBS2950. Additionally, the large reciprocal translocation affects a proline permease (PUT4) regulatory region, resulting in higher PUT4 transcript levels, which agrees with higher ethanol tolerance, improved cell growth when using proline, and higher amino acid consumption during fermentation. These results suggest that SVs are responsible for the rapid physiological adaptation of yeast to an anthropogenic habitat and demonstrate the key contribution of SVs in adaptive fermentative traits in non-model species. ImportanceThe exploration of domestication signatures associated with anthropogenic niches has predominantly focused on studies conducted on model organisms, such as Saccharomyces cerevisiae, overlooking the potential for comparisons across other non-Saccharomyces species. In our research, employing a combination of long- and short-read data, we found domestication signatures in L. cidri, a non-model species recently isolated from fermentative environments in cider in France. The significance of our study lies in the identification of large array of major genomic rearrangements in a cider strain compared to wild isolates, which underly several fermentative traits. These domestication hallmarks result from structural variants, which are likely responsible for the phenotypic differences between strains, providing a rapid path of adaptation to human-related environments.

genetics↗

bioRxiv2022

Natural Variation in Diauxic Shift between Patagonian Saccharomyces eubayanus Strains

The study of natural variation can untap novel alleles with immense value for biotechnological applications. Saccharomyces eubayanus Patagonian isolates exhibit differences in the diauxic shift between glucose and maltose, representing a suitable model to study their natural genetic variation for novel strains for brewing. However, little is known about the genetic variants and chromatin regulators responsible for these differences. Here, we show how genome-wide chromatin accessibility and gene expression differences underlie distinct diauxic shift profiles in S. eubayanus. We identified two strains with a rapid diauxic shift between glucose and maltose (CL467.1 and CBS12357) and one strain with a remarkably low fermentation efficiency and longer lag phase during diauxic shift (QC18). This is associated in the QC18 strain to lower transcriptional activity and chromatin accessibility of specific genes of maltose metabolism, and higher expression levels of glucose transporters. These differences are governed by the HAP complex, which differentially regulates gene expression depending on the genetic background. We found in the QC18 strain a contrasting phenotype to those described in S. cerevisiae, where hap411, hap511 and cin511 knockouts significantly improved the QC18 growth rate in the glucose-maltose shift. The most profound effects were found between CIN5 allelic variants, suggesting that Cin5p could strongly activate a repressor of the diauxic shift in the QC18 strain, but not necessarily in the other strains. The differences between strains could originate from the tree host from which the strains were obtained, which might determine the sugar source preference and the brewing potential of the strain. ImportanceThe diauxic shift has been studied in the budding yeast under laboratory conditions, however, few studies have addressed the diauxic shift between carbon sources under fermentative conditions. Here, we study the transcriptional and chromatin structure differences that explain the natural variation in fermentative capacity and efficiency during diauxic shift of natural isolates of S. eubayanus. Our results show how natural genetic variants in transcription factors impact sugar consumption preferences between strains. These variants have different effects depending on the genetic background, with a contrasting phenotype to those previously described in S. cerevisiae. Our study shows how relatively simple genetic/molecular modifications/editing in the lab can facilitate the study of natural variation of microorganisms for the brewing industry.

microbiology↗