Biology subjects

Liti, G.

Publications and source records attributed to Liti, G..

At least 37 records · Page 2Linked to original sources

bioRxiv2023

Building pangenome graphs

Pangenome graphs can represent all variation between multiple reference genomes, but current approaches to build them exclude complex sequences or are based upon a single reference. In response, we developed the PanGenome Graph Builder (PGGB), a pipeline for constructing pangenome graphs without bias or exclusion. PGGB uses all-to-all alignments to build a variation graph in which we can identify variation, measure conservation, detect recombination events, and infer phylogenetic relationships.

bioinformatics↗

bioRxiv2023

Genome instability footprint under rapamycin and hydroxyurea treatments

The mutational processes dictating the accumulation of mutations in genomes are shaped by genetic background, environment and their interactions. Accurate quantification of mutation rates and spectra under drugs has important implications in disease treatment. Here, we used whole-genome sequencing and time-resolved growth phenotyping of yeast mutation accumulation lines to give a detailed view of the mutagenic effects of rapamycin and hydroxyurea on the genome and cell growth. Mutation rates depended on the genetic backgrounds, but were only marginally affected by rapamycin. As a remarkable exception, rapamycin treatment was associated with frequent chromosome XII amplifications, which compensated for rapamycin induced rDNA repeat contraction on this chromosome and served to maintain rDNA content homeostasis and fitness. In hydroxyurea, a wide range of mutation rates were elevated regardless of the genetic backgrounds, with a particularly high occurrence of aneuploidy that associated with dramatic fitness loss. Hydroxyurea also induced a high T-to-G transversion rate that reversed the common G/C-to-A/T bias in yeast and gave rise to a broad range of structural variants, including mtDNA deletions. The hydroxyurea mutation footprint was consistent with the activation of error-prone DNA polymerase activities and non-homologues end joining repair pathways. Taken together, our study provides an in-depth view of mutation rates and signatures in rapamycin and hydroxyurea and their impact on cell fitness, which brings insights for assessing their chronic effects on genome integrity.

genomics↗

bioRxiv2022

142 telomere-to-telomere assemblies reveal the genome structural landscape in Saccharomyces cerevisiae

As population genomics is transitioning from single reference genomes to pangenomes, major improvements in terms of genome contiguity, phylogenetic sampling, haplotype phasing and structural variant (SV) calling are required. Here, we generated the Saccharomyces cerevisiae Reference Assembly Panel (ScRAP) comprising 142 reference-quality genomes from strains of various geographic and ecological origins that faithfully represent the genomic diversity and complexity of the species. The ca. 4,800 non-redundant SVs we identified impact the expression of genes near the breakpoints and contribute to gene repertoire evolution through disruptions, duplications, fusions and horizontal transfers. We discovered frequent cases of complex aneuploidies, preferentially involving large chromosomes that underwent large SVs. We also characterized the evolutionary dynamics of complex genomic regions that classically remain unassembled in short read-based projects, including the 5 Ty families and the 32 individual telomeres. Overall, the ScRAP represents a crucial step towards establishing a high-quality, unified and complete S. cerevisiae pangenome.

genomics↗

bioRxiv2022

The natural diversity of the yeast proteome reveals chromosome-wide dosage compensation in aneuploids

Aneuploidy, an imbalance in chromosome copy numbers, causes genetic disorders, and drives cancer progression, drug tolerance, and antimicrobial resistance. While aneuploidy can confer stress resistance, it is not well understood how cells overcome the fitness burden caused by aberrant chromosomal copy numbers. Studies using both systematically generated 1-5 and natural aneuploid yeasts 6-8 triggered an intense debate about the role of dosage compensation, concluding that aneuploidy is transmitted to the transcriptome and proteome without significant buffering at the chromosome-wide level, and is, at least in lab strains, associated with significant fitness costs. Conversely, systematic sequencing and phenotyping of large collections of natural isolates revealed that aneuploidy is frequent and has few - if any - fitness costs in nature 9. To address these discrepant findings at the proteomic level, we developed a platform that yields highly precise proteomic measurements across large numbers of genetically diverse samples, and applied it to natural isolates collected as part of the 1011 genomes project 9. For 613 of the isolates, we were able to match the proteomes to their corresponding transcriptomes and genomes, subsequently quantifying the effect of aneuploidy on gene expression by comparing 95 aneuploid with 518 euploid strains. We find, as in previous studies, that aneuploid gene dosage is not buffered chromosome-wide at the transcriptome level. Importantly, in the proteome, we detect an attenuation of aneuploidy by about 25% below the aneuploid gene dosage in natural yeast isolates. Furthermore, this chromosome-wide dosage compensation is associated with the ubiquitin-proteasome system (UPS), which is expressed at higher levels and has increased activity across natural aneuploid strains. Thus, through systematic exploration of the species-wide diversity of the yeast proteome, we shed light on a long-standing debate about the biology of aneuploids, revealing that aneuploidy tolerance is mediated through chromosome-wide dosage compensation at the proteome level.

systems biology↗

bioRxiv2022

Telomeres are shorter in wild Saccharomyces cerevisiae isolates than in domesticated ones

Telomeres are ribonucleoproteins that cap chromosome-ends and their DNA length is controlled by counteracting elongation and shortening processes. The budding yeast Saccharomyces cerevisiae has been a leading model to study telomere DNA length control and dynamics. Its telomeric DNA is maintained at a length that slightly varies between laboratory strains, but little is known about its variation at the species level. The recent publication of the genomes of over 1000 S. cerevisiae strains enabled us to explore telomere DNA length variation at an unprecedented scale. Here, we developed a bioinformatic pipeline (YeaISTY) to estimate telomere DNA length from whole-genome-sequences and applied it to the sequenced 1011 S. cerevisiae collection. Our results revealed broad natural telomere DNA length variation among the isolates. Notably, telomere DNA length is shorter in those derived from wild rather than domesticated environments. Wild isolates are enriched in loss-of-function mutations in genes known to regulate telomere DNA length and the return of domesticated yeasts to a wild habitat coincides with shorter telomeres. Moreover, telomere DNA length variation is associated with mitochondrial metabolism, and this association is driven by wild strains. Overall, these findings suggest that budding yeasts telomere DNA length regulation might be shaped by ecological life-styles.

genomics↗

bioRxiv2022

RecombineX: a generalized computational framework for automatic high-throughput gamete genotyping and tetrad-based recombination analysis

Meiotic recombination is an essential biological process that ensures faithful chromosome segregation and promotes parental allele shuffling. Tetrad analysis is a powerful approach to quantify the genetic makeups and recombination landscapes of meiotic products. Here we present RecombineX (https://github.com/yjx1217/RecombineX), a generalized computational framework that automates the full workflow of marker identification, gamete genotyping, and tetrad-based recombination profiling based on any organisms and genetic backgrounds with batch processing capability. Aside from conventional reference-based analysis, RecombineX can also perform analysis based on parental genome assemblies, which enables analyzing meiotic recombination landscapes in their native genomic contexts. Additional features such as copy number variation profiling and missing genotype inference further enhance downstream analysis. RecombineX also includes a dedicate module for simulating the genomes and reads of recombinant tetrads, which enables fine-tuned simulation-based hypothesis testing. This simulation module revealed the power and accuracy of RecombineX even when analyzing tetrads with very low sequencing depths (e.g., 1-2X). Tetrad sequencing data from the budding yeast Saccharomyces cerevisiae and green alga Chlamydomonas reinhardtii were further used to demonstrate the accuracy and robustness of RecombineX for organisms with both small and large genomes, manifesting RecombineX as an all-around one stop solution for future tetrad analysis. Author SummaryMeiosis is a fundamental cellular process that ensures faithful chromosome segregation and promotes allele shuffling. Tetrad analysis, which isolates and genotypes all four meiotic products (i.e., tetrad) derived from a single meiosis, remains the most straightforward and powerful way of studying meiotic recombination and its modulators at fine scales. The wide application of tetrad analysis in yeasts, filamentous fungi, green algae, and land plants have substantially expand our understanding of meiotic recombination in terms of both genome-wide landscapes and molecular mechanisms. Here we described the first generalized computational framework named RecombineX that automates the full workflow of tetrad analysis based on any organisms and genetic backgrounds. In addition, aside from conventional reference-based analysis, RecombineX can also perform analysis based on parental genome assemblies, which enables analyzing meiotic recombination landscapes in their native genomic contexts. Using both simulated and real tetradsequencing data, we further demonstrated RecombineXs trustable performance, versatile usage, and batch-processing capability, manifesting RecombineX as an all-around one stop solution for tetrad analysis. Especially considering that meiotic gamete genome sequencing from different natural and mutant backgrounds can now be acquired, we expect RecombineX to become a popular tool that empowers future tetrad analysis across different genetic backgrounds and species.

bioinformatics↗

bioRxiv2021

Unlocking the functional potential of polyploid yeasts

Breeding and domestication have generated widely exploited crops, animals and microbes. However, many Saccharomyces cerevisiae industrial strains have complex polyploid genomes and are sterile, preventing genetic improvement strategies based on breeding. Here, we present a novel strain improvement approach based on the budding yeasts property to promote genetic recombination when meiosis is interrupted and cells return-to-mitotic-growth (RTG). We demonstrated that two unrelated sterile industrial strains with complex triploid and tetraploid genomes were RTG-competent and developed a visual screening for easy and high-throughput identification of recombined RTG clones based on colony phenotypes. Sequencing of the evolved clones revealed unprecedented levels of RTG-induced genome-wide recombination. We generated and extensively phenotyped a RTG library and identified clones with superior biotechnological traits. Thus, we propose the RTG-framework as a fully non-GMO workflow to rapidly improve industrial yeasts that can be easily brought to the market.

genetics↗

bioRxiv2021

Deep sampling of ancestral genetic diversity reveals Saccharomyces cerevisiae pre-domestication life histories

The ecology and genetic diversity of model yeast Saccharomyces cerevisiae prior to human domestication remain poorly understood. Taiwan is regarded as part of this yeasts geographic birthplace where the most divergent natural lineage was discovered. Here, we deep sampled the broad-leaf forests across this continental island to probe the ancestral species diversity. We found that S. cerevisiae is distributed ubiquitously at low abundance in the forests. Whole-genome sequencing of 121 isolates revealed nine distinct lineages, the highest known in any region. Three lineages are endemic to Taiwan and six are widespread in Asia. Molecular dating placed the divergence of the Taiwanese and Asian lineages during the Pleistocene, when a transient continental shelf land bridge connected Taiwan to other major landmasses. Extensive historical and recent admixture events were detected between natural lineages. In particular, the genetic component from a lineage associated with fruits that spanned the widest geographical range was present in most admixed isolates. Collectively, Taiwanese isolates harbor genetic diversity comparable to that of the whole Asia continent, and different lineages have coexisted at a fine spatial scale even on the same tree. Patterns of variations within each lineage revealed that S. cerevisiae is highly clonal and predominantly reproduces asexually in nature. We detected prevalent purifying selection genome-wide, with lineage-specific signals of positive or directional selection independent between lineages. This study establishes that S. cerevisiae has rich natural diversity sheltered from human influences, making it a powerful model system in microbial ecology.

genomics↗

bioRxiv2021

Lager yeast design through meiotic segregation of a fertile Saccharomyces cerevisiae x Saccharomyces eubayanus hybrid

Yeasts in the lager brewing group are closely related and consequently do not exhibit significant genetic variability. Here, an artificial Saccharomyces cerevisiae x Saccharomyces eubayanus tetraploid interspecies hybrid was created by rare mating, and its ability to sporulate and produce viable gametes was exploited to generate phenotypic diversity. Four spore clones obtained from a single ascus were isolated, and their brewing-relevant phenotypes were assessed. These F1 spore clones were found to differ with respect to fermentation performance under lager brewing conditions (15 {degrees}C, 15 {degrees}Plato), production of volatile aroma compounds, flocculation potential and temperature tolerance. One spore clone, selected for its rapid fermentation and acetate ester production was sporulated to produce an F2 generation, again comprised of four spore clones from a single ascus. Again, phenotypic diversity was introduced. In two of these F2 clones, the fermentation performance was maintained and acetate ester production was improved relative to the F1 parent and the original hybrid strain. Strains also performed well in comparison to a commercial lager yeast strain. Spore clones varied in ploidy and chromosome copy numbers, and faster wort fermentation was observed in strains with a higher ploidy. An F2 spore clone was also subjected to 10 consecutive wort fermentations, and single cells were isolated from the resulting yeast slurry. These isolates also exhibited variable fermentation performance and chromosome copy numbers, highlighting the instability of polyploid interspecific hybrids. These results demonstrate the value of this natural approach to increase the phenotypic diversity of lager brewing yeast strains. Contribution to the fieldLager beer fermentations have traditionally been carried out with natural S. cerevisiae x S. eubayanus hybrids. These strains possess both the ability to tolerate low temperatures and the ability to utilize efficiently wort sugars. However, being closely related, strains within the group exhibit limited phenotypic variability. Since the recent discovery of wild strains of S. eubayanus, it has been possible to generate lager yeast hybrids artificially, thereby increasing the genetic and phenotypic diversity of lager brewing strains. Here, to demonstrate the potential for further increased diversity, a constructed tetraploid hybrid was sporulated and spore clones derived from a single ascus were evaluated with respect to fermentation performance (sugar utilization, stress tolerance and volatile aroma synthesis). Meiosis introduced variability in a number of key parameters. One fertile spore clone from this F1 generation was sporulated to introduce further diversity and to demonstrate the potential of clone selection in steering phenotypes in a desirable direction. Genome instability of hybrids was observed, but this can be exploited to further increase diversity. This was demonstrated by assessing performance of variants isolated after ten consecutive rounds of fermentation. The approach allows for the introduction of phenotypic diversity without the need for targeted genetic modification.

microbiology↗

bioRxiv2020

Aborting meiosis overcomes hybrid sterility

Hybrids between species or diverged lineages contain fundamentally novel genetic combinations but an impaired meiosis often makes them evolutionary dead ends. Here, we explored to what extent and how an aborted meiosis followed by a return-to-growth (RTG) promotes recombination across a panel of 20 yeast diploid backgrounds with different genomic structures and levels of sterility. Genome analyses of 284 clones revealed that RTG promoted recombination and generated extensive regions of loss-of-heterozygosity in sterile hybrids with either a defective meiosis or a heavily rearranged karyotype, whereas RTG recombination was reduced by high sequence divergence between parental subgenomes. The RTG recombination preferentially occurred in regions with local sequence homology and in meiotic recombination hotspots. The loss-of-heterozygosity had a profound impact on sexual and asexual fitness, and enabled genetic mapping of phenotypic differences in sterile lineages where linkage or association analyses failed. We propose that RTG gives sterile hybrids access to a natural route for genome recombination and adaptation. One sentence summaryAborting meiosis followed by a return to mitotic growth promotes evolution by genome wide-recombination in sterile yeast hybrids.

genetics↗

bioRxiv2020

Diverse natural variants suppress mutations in hundreds of essential genes

The consequence of a mutation can be influenced by the context in which it operates. For example, loss of gene function may be tolerated in one genetic background, but lead to lethality in another. The extent to which mutant phenotypes are malleable, the complexity of the architecture of modifiers, and the identities of causal genes and pathways remain largely unknown. Here, we measure the fitness effects of ~1,500 temperature sensitive alleles of yeast essential genes in the context of variation from ten different natural genetic backgrounds, and map the modifiers for 19 combinations. Altogether, fitness defects for 183 of the 530 tested genes (35%) could be suppressed by standing genetic variation in at least one wild strain. Suppression was generally driven by gain-of-function of a single, strong modifier gene. The validated causes included both variants in protein interaction partners or pathway members suppressing specific genes, as well as general modifiers altering the effect of many temperature sensitive alleles. The emerging frequency of suppression and range of possible suppression mechanisms suggest that a substantial fraction of monogenic diseases could be repressed by modulating other gene products.

genetics↗

bioRxiv2020

Control of mitochondrial superoxide production includes programmed mtDNA deletion and restoration

Deletion of mitochondrial DNA in eukaryotes is mainly attributed to rare accidental events associated with mitochondrial replication or repair of double-strand breaks. We report the discovery that yeast cells arrest harmful intramitochondrial superoxide production by shutting down respiration through genetically controlled deletion of mitochondrial oxidative phosphorylation genes. We show that the regulatory circuitry underlying this editing critically involves the antioxidant enzyme superoxide dismutase 2 and two-way mitochondrial-nuclear communication. While mitochondrial DNA homeostasis is rapidly restored after cessation of a short-term superoxide stress, long-term stress causes maladaptive persistence of the deletion process, leading to complete annihilation of the cellular pool of intact mitochondrial genomes and irrevocable loss of respiratory ability. Our results may therefore be of etiological as well as therapeutic importance with regard to age-related mitochondrial impairment and disease. One-Sentence SummaryGenetically controlled editing of mitochondrial DNA is an integral part of the yeasts defenses against oxidative damage.

cell biology↗

bioRxiv2020

CRISpy-pop: a web tool for designing CRISPR/Cas9-driven genetic modifications in diverse populations

CRISPR/Cas9 is a powerful tool for editing genomes, but design decisions are generally made with respect to a single reference genome. With population genomic data becoming available for an increasing number of model organisms, researchers are interested in manipulating multiple strains and lines. CRISpy-pop is a web application that generates and filters guide RNA sequences for CRISPR/Cas9 genome editing for diverse yeast and bacterial strains. The current implementation designs and predicts the activity of guide RNAs against more than 1000 Saccharomyces cerevisiae genomes, including 167 strains frequently used in bioenergy research. Zymomonas mobilis, an increasingly popular bacterial bioenergy research model, is also supported. CRISpy-pop is available as a web application (https://CRISpy-pop.glbrc.org/) with an intuitive graphical user interface. CRISpy-pop also cross-references the human genome to allow users to avoid the selection of sgRNAs with potential biosafety concerns. Additionally, CRISpy-pop predicts the strain coverage of each guide RNA within the supported strain sets, which aids in functional population genetic studies. Finally, we validate how CRISpy-pop can accurately predict the activity of guide RNAs across strains using population genomic data.

genetics↗

bioRxiv2020

Domestication reprogrammed the budding yeast life cycle

Domestication of plants and animals is the foundation for feeding the world population. We report that domestication of the model yeast S. cerevisiae reprogrammed its life cycle entirely. We tracked growth, gamete formation and cell survival across many environments for nearly 1000 genome sequenced isolates and found a remarkable dichotomy between domesticated and wild yeasts. Wild yeasts near uniformly trigger meiosis and sporulate when encountering nutrient depletions, whereas domestication relaxed selection on sexual reproduction and favoured survival as quiescent cells. Domestication also systematically enhanced fermentative over respiratory traits while decreasing stress tolerance. We show that this yeast domestication syndrome was driven by aneuploidies and gene function losses that emerged independently in multiple domesticated lineages during the species recent evolutionary history. We found domestication to be the most dramatic event in budding yeast evolution, raising questions on how much domestication has distorted our understanding of this key model species.

evolutionary biology↗

bioRxiv2019

Discordant evolution of mitochondrial and nuclear yeast genomes at population level

Mitochondria are essential organelles partially regulated by their own genomes. The mitochondrial genome maintenance and inheritance differ from nuclear genome, potentially uncoupling their evolutionary trajectories. Here, we analysed mitochondrial sequences obtained from the 1,011 Saccharomyces cerevisiae strain collection and identified pronounced differences with their nuclear genome counterparts. In contrast with most fungal species, S. cerevisiae mitochondrial genomes show higher genetic diversity compared to the nuclear genomes. Strikingly, mitochondrial genomes appear to be highly admixed, resulting in a complex interconnected phylogeny with weak grouping of isolates, whereas interspecies introgressions are very rare. Complete genome assemblies revealed that structural rearrangements are nearly absent with rare inversions detected. We tracked introns variation in COX1 and COB to infer gain and loss events throughout the species evolutionary history. Mitochondrial genome copy number is connected with the nuclear genome and linearly scale up with ploidy. We observed rare cases of naturally occurring mitochondrial DNA loss, petite, with a subset of them that do not suffer fitness growth defects. Overall, our results illustrate how differences in the biology of two genomes coexisting in the same cells can lead to discordant evolutionary histories.

genomics↗

bioRxiv2019

An Out-of-Patagonia dispersal explains most of the worldwide genetic distribution in Saccharomyces eubayanus

Saccharomyces eubayanus represents missing cryotolerant ancestor of lager yeast hybrid and can be found in Patagonia in association with Nothofagus forests. The limited number of isolates and associated genomes available has prevented to resolve the S. eubayanus origin and evolution. Here, we present a sampling effort at an unprecedented scale and report the isolation of 160 strains from ten sampling sites along 2,000 km distance in South America. We sequenced the genome of 82 strains and, together with other 25 available genomes, performed comprehensive phylogenetic analysis. Our results revealed the presence of three main Patagonia-B lineages together with dozens of admixed strains distributed in three mosaic clusters. The PB-1 lineage isolated from Tierra del Fuego exhibited the highest genetic diversity, lowest LD blocks and highest Fis values compared to the other lineages, suggesting a successful adaptation to cold temperatures in extreme environments and greater inbreeding rates in Tierra del Fuego. Differences between lineages and strains were found in terms of aneuploidy and pangenome content, evidencing a lateral gene transfer event in PB-2 strains from an unknown donor species. Overall, the Patagonian lineages, particularly southern populations, showed a greater global genetic diversity compared to Holarctic and Chinese lineages, supporting the scenario of a S. eubayanus colonization from Patagonia and then spread towards northern and western regions, including the Holarctic (North America and China) and New Zealand. Interestingly, fermentative capacity and maltose consumption resulted negatively correlated with latitude, indicating a better fermentative performance in norther populations. Our genome analysis together with previous reports in the sister species S. uvarum strongly suggests that the S. eubayanus ancestor could have originated in Patagonia or the Southern Hemisphere, rather than China, yet further studies are needed to resolve this conflicting scenario. Understanding S. eubayanus evolutionary history is crucial to resolve the unknown origin of the lager yeast and might open new avenues for biotechnological applications.

genomics↗

bioRxiv2019

Intragenic repeat expansions control yeast chronological aging

Aging varies among individuals due to both genetics and environment but the underlying molecular mechanisms remain largely unknown. Using a highly recombined Saccharomyces cerevisiae population, we found 30 distinct Quantitative Trait Loci (QTLs) that control chronological life span (CLS) in calorie rich and calorie restricted environments, and under rapamycin exposure. Calorie restriction and rapamycin extended life span in virtually all genotypes, but through different genetic variants. We tracked the two major QTLs to massive expansions of intragenic tandem repeats in the cell wall glycoproteins FLO11 and HPF1, which caused a dramatic life span shortening. Life span impairment by N-terminal HPF1 repeat expansion was partially buffered by rapamycin but not by calorie restriction. The HPF1 repeat expansion shifted yeast cells from a sedentary to a buoyant state, thereby increasing their exposure to surrounding oxygen. The higher oxygenation perturbed methionine, lipid, and purine metabolism, which likely explains the life span shortening. We conclude that fast evolving intragenic repeat expansions can fundamentally change the relationship between cells and their environment with profound effects on cellular life style and longevity.

genomics↗

bioRxiv2018

Accurate tracking of the mutational landscape of diploid hybrid genomes reveals genetic background effects

BackgroundGenome evolution promotes diversity within a population via mutations, recombination, and whole-genome duplication. However, quantifying precisely these factors in diploid hybrid genomes is challenging. Here we present an integrated experimental and computational workflow to accurately track the mutational landscape of yeast diploid hybrids (MuLoYDH) in terms of single-nucleotide variants, small insertions/deletions, copy-number variants and loss-of-heterozygosity. ResultsHaploid Saccharomyces parents were combined into diploid hybrids with fully phased genome and controlled levels of heterozygosity. The resulting hybrids represented the ancestral state and were evolved under different laboratory protocols. Variant simulations enabled to efficiently integrate competitive and standard mapping, depending on local levels of heterozygosity and read length. Experimental validations proved high accuracy and resolution of our computational approach. Finally, applying MuLoYDH to four different diploids revealed striking genetic background effects. Homozygous S. cerevisiae showed ~4-fold higher mutation rate compared to S. paradoxus. In contrast, interspecies hybrids exhibited mutation rates similar to intraspecies hybrids despite 10-fold higher heterozygosity. MuLoYDH unveiled that a substantial fraction of the genome (~200 bp per generation) was shaped by loss-of-heterozygosity and this process was strongly inhibited by high levels of heterozygosity. ConclusionsWe report a comprehensive framework for characterizing the mutational spectrum of yeast diploid hybrids with unprecedented resolution, which can be generalised to other genetic systems. Applying MuLoYDH to laboratory-evolved hybrids provided novel quantitative insights into the evolutionary processes that mould yeast genomes.

genomics↗