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At least 163 records · Page 9

Barcoded overexpression screens in gut Bacteroidales identify genes with roles in carbon utilization and stress resistance

Abstract A mechanistic understanding of host-microbe interactions in the gut microbiome is hindered by poorly annotated bacterial genomes. While functional genomics can generate large gene-to-phenotype datasets to accelerate functional discovery, their applications to study gut anaerobes have been limited. For instance, most gain-of-function screens of gut-derived genes have been performed in Escherichia coli and assayed in a small number of conditions. To address these challenges, we develop Barcoded Overexpression BActerial shotgun library sequencing (Boba-seq). We demonstrate the power of this approach by assaying genes from diverse gut Bacteroidales overexpressed in Bacteroides thetaiotaomicron . From hundreds of experiments, we identify new functions and phenotypes for 29 genes important for carbohydrate metabolism or tolerance to antibiotics or bile salts. Highlights include the discovery of a d -glucosamine kinase, a raffinose transporter, and several routes that increase tolerance to ceftriaxone and bile salts through lipid biosynthesis. This approach can be readily applied to develop screens in other strains and additional phenotypic assays.

59 BASIC BIOLOGICAL SCIENCES↗

Genome shuffling enables quantitative trait locus mapping in Bacillus subtilis

Genetic mapping is a powerful tool for eukaryotic genetics that has only been applied to bacteria in limited circumstances. Quantitative trait locus (QTL) mapping generally relies on sexual recombination to break linkages between genes, yet bacteria rarely undergo sufficient homologous recombination to generate suitable mapping populations. In this work, we used iterative biparental genome shuffling by protoplast fusion in Bacillus subtilis to generate a population of bacteria with substantial random recombination throughout their genomes. Individual shuffled progeny were arrayed in well plates, resequenced, and characterized for a range of complex phenotypes including spore germination and swarming motility. Genetic mapping of the resulting phenotypes identified high-confidence QTLs of moderate size (~10 kb), and these associations were validated through targeted genetic swaps. This B. subtilis QTL population can easily be used to map additional phenotypes, and the general approach for QTL mapping is applicable in a wide range of bacteria.

Vasileva, Delyana [ORNL] (ORCID:0000000279197883)↗

Computer vision models enable mixed linear modeling to predict arbuscular mycorrhizal fungal colonization using fungal morphology

Abstract The presence of Arbuscular Mycorrhizal Fungi (AMF) in vascular land plant roots is one of the most ancient of symbioses supporting nitrogen and phosphorus exchange for photosynthetically derived carbon. Here we provide a multi-scale modeling approach to predict AMF colonization of a worldwide crop from a Recombinant Inbred Line (RIL) population derived from Sorghum bicolor and S. propinquum . The high-throughput phenotyping methods of fungal structures here rely on a Mask Region-based Convolutional Neural Network (Mask R-CNN) in computer vision for pixel-wise fungal structure segmentations and mixed linear models to explore the relations of AMF colonization, root niche, and fungal structure allocation. Models proposed capture over 95% of the variation in AMF colonization as a function of root niche and relative abundance of fungal structures in each plant. Arbuscule allocation is a significant predictor of AMF colonization among sibling plants. Arbuscules and extraradical hyphae implicated in nutrient exchange predict highest AMF colonization in the top root section. Our work demonstrates that deep learning can be used by the community for the high-throughput phenotyping of AMF in plant roots. Mixed linear modeling provides a framework for testing hypotheses about AMF colonization phenotypes as a function of root niche and fungal structure allocations.

59 BASIC BIOLOGICAL SCIENCES↗

Intraspecific Reaction Norm Variation Controls the Eco-Evolutionary Consequences of Environmental Change

As environmental change accelerates globally, understanding concurrent organismal, species, and community responses is increasingly vital. Here, we examine these collective responses by incorporating genotype-specific thermal reaction norms into an eco-evolutionary predator-prey model, allowing us to track simultaneous phenotypic, ecological, and evolutionary responses to environmental change within ecological communities. We show that the reaction norms expressed by genotypes within a population determine how a community switches between different eco-evolutionary outcomes with changes in temperature. We identify how different components of phenotypic variation in thermal reaction norms—environmental (E), additive environmental and genetic (E + G), and gene-by-environment interactions (G × E)—influence eco-evolutionary dynamics and outcomes as temperature changes. Furthermore, our findings underscore how complex eco-evolutionary responses to environmental change ultimately emerge from variation in reaction norms among genotypes, offering new mechanistic insights into environmental impacts on adaptation, the maintenance of phenotypic and genetic variation, and ecological stability, which is crucial for understanding and predicting eco-evolutionary effects of rapid environmental change in the future.

Eco-evolutionary↗

Induced and natural variation affect traits independently in hybrid Populus

Abstract The genetic control of many plant traits can be highly complex. Both allelic variation (sequence change) and dosage variation (copy number change) contribute to a plant's phenotype. While numerous studies have investigated the effect of allelic or dosage variation, very few have documented both within the same system, leaving their relative contribution to phenotypic effects unclear. The Populus genome is highly polymorphic, and poplars are fairly tolerant of gene dosage variation. Here, using a previously established Populus hybrid F1 population, we assessed and compared the effect of natural allelic variation and induced dosage variation on biomass, phenology, and leaf morphology traits. We identified QTLs for many of these traits, but our results indicate limited overlap between the QTLs associated with natural allelic variation and induced dosage variation. Additionally, the integration of data from both allelic and dosage variation identifies a larger set of QTLs that together explain a larger percentage of the phenotypic variance. Finally, our results suggest that the effect of the large indels might mask that of allelic QTLs. Our study helps clarify the relationship between allelic and dosage variation and their effects on quantitative traits.

Guo, Weier (ORCID:0000000251789334)↗

Bacterial and fungal composition and exometabolites control the development and persistence of soil water repellency

Soil water repellency (SWR), the reduced affinity of soil for water, is a phenomenon that affects soils globally. With worsening climate change, SWR is expected to increase emphasizing the need to understand the mechanisms driving SWR development and persistence. The importance of the soil microbes in SWR has been postulated for decades, but limited research has been conducted into whole-community interactions and the role of community metabolic activity. To address this gap in knowledge, we investigated the direct effect of microbial community composition, activity, and diversity, as well as their associated metabolites on the development and persistence of SWR by inoculating microcosms containing model soils with 15 different microbial communities and quantified respiration and SWR over time. Six communities that consistently produced either a hydrophobic or hydrophilic phenotype were characterized using metagenomics and metabolomics to determine the impact of microbial and metabolite composition and diversity on SWR. We identified several bacterial genera with significant changes in abundance between SWR phenotypes including Nocardiopsis and Kocuria in hydrophilic and Streptomyces and Cutibacterium in hydrophobic. We discovered that hydrophilic communities were more positively connected when compared to hydrophobic communities, which could be due to an increase in defense mechanism genes. Additionally, we identified specific metabolites associated with hydrophilic and hydrophobic phenotypes including an increase in the osmolyte ectoine in hydrophilic and an increase in plant-derived decomposition products in hydrophobic communities. Finally, our research suggests that fungi, previously thought to cause hydrophobicity, may actually contribute to hydrophilicity through their preferential consumption of hydrophobic compounds.

54 ENVIRONMENTAL SCIENCES↗

Breaking the barrier of human-annotated training data for machine learning-aided plant research using aerial imagery

Machine learning (ML) can accelerate biological research. However, the adoption of such tools to facilitate phenotyping based on sensor data has been limited by (i) the need for a large amount of human-annotated training data for each context in which the tool is used and (ii) phenotypes varying across contexts defined in terms of genetics and environment. This is a major bottleneck because acquiring training data is generally costly and time-consuming. This study demonstrates how a ML approach can address these challenges by minimizing the amount of human supervision needed for tool building. A case study was performed to compare ML approaches that examine images collected by an uncrewed aerial vehicle to determine the presence/absence of panicles (i.e. “heading”) across thousands of field plots containing genetically diverse breeding populations of 2 Miscanthus species. Automated analysis of aerial imagery enabled the identification of heading approximately 9 times faster than in-field visual inspection by humans. Leveraging an Efficiently Supervised Generative Adversarial Network (ESGAN) learning strategy reduced the requirement for human-annotated data by 1 to 2 orders of magnitude compared to traditional, fully supervised learning approaches. The ESGAN model learned the salient features of the data set by using thousands of unlabeled images to inform the discriminative ability of a classifier so that it required minimal human-labeled training data. This method can accelerate the phenotyping of heading date as a measure of flowering time in Miscanthus across diverse contexts (e.g. in multistate trials) and opens avenues to promote the broad adoption of ML tools.

59 BASIC BIOLOGICAL SCIENCES↗

Rhizobial Secretion of Truncated Exopolysaccharides Severely Impairs the Mesorhizobium-Lotus Symbiosis

The symbiosis between Mesorhizobium japonicum R7A and Lotus japonicus Gifu is an important model system for investigating the role of bacterial exopolysaccharides (EPS) in plant-microbe interactions. Previously, we showed that R7A exoB mutants that are affected at an early stage of EPS synthesis and in lipopolysaccharide (LPS) synthesis induce effective nodules on L. japonicus Gifu after a delay, whereas exoU mutants affected in the biosynthesis of the EPS side chain induce small uninfected nodule primordia and are impaired in infection. The presence of a halo around the exoU mutant when grown on Calcofluor-containing media suggested the mutant secreted a truncated version of R7A EPS. A nonpolar ΔexoA mutant defective in the addition of the first glucose residue to the EPS backbone was also severely impaired symbiotically. Here, we used a suppressor screen to show that the severe symbiotic phenotype of the exoU mutant was due to the secretion of an acetylated pentasaccharide, as both monomers and oligomers, by the same Wzx/Wzy system that transports wild-type exopolysaccharide. We also present evidence that the ΔexoA mutant secretes an oligosaccharide by the same transport system, contributing to its symbiotic phenotype. In contrast, ΔexoYF and polar exoA and exoL mutants have a similar phenotype to exoB mutants, forming effective nodules after a delay. These studies provide substantial evidence that secreted incompatible EPS is perceived by the plant, leading to abrogation of the infection process.

59 BASIC BIOLOGICAL SCIENCES↗

Hidden Allies: Decoding the Core Endohyphal Bacteriome of Aspergillus fumigatus

ABSTRACT Bacterial–fungal interactions that influence the behaviour of one or both organisms are common in nature. Well‐studied systems include endosymbiotic relationships that range from transient to long‐term associations. Diverse endohyphal bacteria associate with fungal hosts, emphasising the need to better comprehend the fungal bacteriome. We evaluated the hypothesis thatAspergillus fumigatusharbours an endohyphal community of bacteria that influence the host phenotype. We analysed whether 38A. fumigatusstrains show stable association with diverse endohyphal bacteria; all derived from single‐conidium cultures that were subjected to antibiotic and heat treatments. The fungal bacteriome, inferred through analysis of bacterial diversity within the fungal strains (short‐ and long‐ read sequencing methods), revealed the presence of core endohyphal bacterial genera. Microscopic analysis further confirmed the presence of endohyphal bacteria. The fungal strains exhibited high genetic diversity and phenotypic heterogeneity in drug susceptibility and in vivo virulence. No correlations were observed between genomic or functional traits and bacteriome diversity, but the abundance of some bacterial genera correlated with fungal virulence or posaconazole susceptibility. The observed endobacteriome may play functional roles, for example, nitrogen fixation. Our study emphasises the existence of complex interactions between fungi and endohyphal bacteria, possibly impacting the phenotype of the fungal host, including virulence.

Environmental Sciences & Ecology↗

A small number of point mutations confer formate tolerance in Shewanella oneidensis

ABSTRACT Microbial electrosynthesis (MES) is a sustainable approach to chemical production from CO 2 and clean electricity. However, limitations in electron transfer efficiency and gaps in understanding of electron transfer pathways in MES systems prevent full realization of this technology. Shewanella oneidensis could serve as an MES biocatalyst because it has a well-studied, efficient transmembrane electron transfer pathway. A key first step in MES in this organism could be CO 2 reduction to formate. However, we report that wild-type S. oneidensis does not tolerate high levels of formate. In this work, we created and characterized formate-tolerant strains of S. oneidensis for further engineering and future use in MES systems through adaptive laboratory evolution. Two different point mutations in a gene encoding a predicted sodium-dependent bicarbonate transporter and a DUF2721-containing protein separately confer formate tolerance to S. oneidensis . The mutations were further evaluated to understand their role in improving formate tolerance. We also show that the wild-type and mutant versions of the putative sodium-dependent bicarbonate transporter improve formate tolerance of Zymomonas mobilis , indicating the potential of transferring this formate tolerance phenotype to other organisms. IMPORTANCE Shewanella oneidensis is a bacterium with a well-studied, efficient extracellular electron transfer pathway. This capability could make this organism a suitable host for microbial electrosynthesis using CO 2 or formate as feedstocks. However, we report here that formate is toxic to S. oneidensis , limiting the potential for its use in these systems. In this work, we evolve several strains of S. oneidensis that have improved formate tolerance, and we investigate some mutations that confer this phenotype. The phenotype is confirmed to be attributed to several single point mutations by transferring the wild-type and mutant versions of each gene to the wild-type strain. Finally, the formate tolerance mechanism of one variant is studied using structural modeling and expression in another host. This study, therefore, presents a simple method for conferring formate tolerance to bacterial hosts.

Cross, Megan C. Gruenberg (ORCID:0000000291589900)↗

Alternate routes to acetate tolerance lead to varied isoprenol production from mixed carbon sources in Pseudomonas putida

ABSTRACT Lignocellulose is a renewable resource for the production of a diverse array of platform chemicals, including the biofuel isoprenol. Although this carbon stream provides a rich source of sugars, other organic compounds, such as acetate, can be used by microbial hosts. Here, we examined the growth and isoprenol production in a Pseudomonas putida strain pre-tolerized (“PT”) background where its native isoprenol catabolism pathway is deleted, using glucose and acetate as carbon sources. We found that PT displays impaired growth in minimal medium containing acetate and often fails to grow in glucose-acetate medium. Using a mutant recovery-based approach, we generated tolerized strains that overcame these limitations, achieving fast growth and isoprenol production in the mixed carbon feed. Changes in the glucose and acetate assimilation routes, including an upregulation in PP_0154 (SpcC, succinyl-CoA:acetate CoA-transferase) and differential expression of the gluconate assimilation pathways, were key for higher isoprenol titers in the tolerized strains, whereas a different set of mechanisms were likely enabling tolerance phenotypes in media containing acetate. Among these, a coproporphyrinogen-III oxidase (HemN) was upregulated across all tolerized strains and in one isolate required for acetate tolerance. Utilizing a defined glucose and acetate mixture ratio reflective of lignocellulosic feedstocks for isoprenol production in P. putida allowed us to obtain insights into the dynamics and challenges unique to dual carbon source utilization that are obscured when studied separately. Together, this enabled the development of a P. putida bioconversion chassis able to use a more complex carbon stream to produce isoprenol. IMPORTANCE Acetate is a relatively abundant component of many lignocellulosic carbon streams and has the potential to be used together with sugars, especially in microbes with versatile catabolism such as P. putida . However, the use of mixed carbon streams necessitates additional optimization. Furthermore, the use of P. putida for the production of the biofuel target, isoprenol, requires the use of engineered strains that have additional growth and production constraints when cultivated in acetate and glucose mixtures. In this study, we generate acetate-tolerant P. putida strains that overcome these challenges and examine their ability to produce isoprenol. We show that acetate tolerance and isoprenol production, although independent phenotypes, can both be optimized in a given P. putida strain. Using proteomics and whole genome sequencing, we examine the molecular basis of both phenotypes and show that tolerance to acetate can occur via alternate routes and result in different impacts on isoprenol production.

de Siqueira, Guilherme M. V. (ORCID:00000002364563↗

Catabolic pathway acquisition by rhizosphere bacteria readily enables growth with a root exudate component but does not affect root colonization

Horizontal gene transfer (HGT) is a fundamental evolutionary process that plays a key role in bacterial evolution. The likelihood of a successful transfer event is expected to depend on the precise balance of costs and benefits resulting from pathway acquisition. Most experimental analyses of HGT have focused on phenotypes that have large fitness benefits under appropriate selective conditions, such as antibiotic resistance. However, many examples of HGT involve phenotypes that are predicted to provide smaller benefits, such as the ability to catabolize additional carbon sources. We have experimentally simulated the consequences of one such HGT event in the laboratory, studying the effects of transferring a pathway for catabolism of the plant-derived aromatic compound salicyl alcohol between rhizosphere isolates from the Pseudomonas genus. We find that pathway acquisition enables rapid catabolism of salicyl alcohol with only minor disruptions to the existing metabolic and regulatory networks of the new host. However, this new catabolic potential does not confer a measurable fitness advantage during competitive growth in the rhizosphere. We conclude that the phenotype of salicyl alcohol catabolism is readily transferable but is selectively neutral under environmentally relevant conditions. We propose that this condition is common and that HGT of many pathways will be self-limiting because the selective benefits are small.

59 BASIC BIOLOGICAL SCIENCES↗

Multi-strain analysis of Pseudomonas putida reveals the metabolic and genetic diversity of the species

Pseudomonas putida is a gram-negative bacterial species increasingly utilized in biotechnology due to its robust growth, ability to degrade aromatic compounds, solvent tolerance, and genetic tractability. In this study, we report a comprehensive multi-strain analysis of 164 P. putida strains based on the reconstruction of a pan-putida metabolic network and the formulation of strain-specific genome-scale metabolic models (GEMs). We performed whole-genome sequencing and hybrid assembly for 40 strains, contributing a ~8% increase to the available genomic data for P. putida . Furthermore, high-throughput phenotypic profiling using the Biolog phenotype microarray system for 24 strains on 190 unique carbon sources, along with 15 aromatic compounds not present on Biolog plates, yielded 4,920 unique strain-phenotype measurements. These data were leveraged to curate GEMs for 24 representative strains, including a refined model for strain KT2440, which comprised 1,480 genes and 2,191 metabolites, achieving a prediction accuracy of 91.2% in carbon utilization. Systematic comparison of genomes and GEMs revealed both conserved core pathways and significant allelic and functional divergence across strains, highlighting strain-specific variation in aromatic degradation. While pathways for protocatechuate and phenylacetate degradation were widely conserved, metabolic capabilities for compounds such as ferulate, phenol, and cresols varied markedly, suggesting adaptation to distinct ecological niches. Alleleome analysis of enzymes, such as PcaI and PcaJ, revealed distinct, functionally similar clades, indicating possible convergent evolution or horizontal gene transfer. These results provide computable resources and informative models for selecting P. putida strains with desired traits for biomanufacturing and bioremediation and offer insights into the evolution and phylogeny of the P. putida species.

aromatics utilization↗

pixelvar79/ESGAN-Flowering-Detection-paper

Machine learning (ML) can accelerate biological research. However, the adoption of such tools to facilitate phenotyping based on sensor data has been limited by (i) the need for a large amount of human-annotated training data for each context in which the tool is used and (ii) phenotypes varying across contexts defined in terms of genetics and environment. This is a major bottleneck because acquiring training data is generally costly and time-consuming. This study demonstrates how a ML approach can address these challenges by minimizing the amount of human supervision needed for tool building. A case study was performed to compare ML approaches that examine images collected by an uncrewed aerial vehicle to determine the presence/absence of panicles (i.e. “heading”) across thousands of field plots containing genetically diverse breeding populations of 2 Miscanthus species. Automated analysis of aerial imagery enabled the identification of heading approximately 9 times faster than in-field visual inspection by humans. Leveraging an Efficiently Supervised Generative Adversarial Network (ESGAN) learning strategy reduced the requirement for human-annotated data by 1 to 2 orders of magnitude compared to traditional, fully supervised learning approaches. The ESGAN model learned the salient features of the data set by using thousands of unlabeled images to inform the discriminative ability of a classifier so that it required minimal human-labeled training data. This method can accelerate the phenotyping of heading date as a measure of flowering time in Miscanthus across diverse contexts (e.g. in multistate trials) and opens avenues to promote the broad adoption of ML tools.

Varela, Sebastian↗

Nitrogen starvation causes lipid remodeling in Rhodotorula toruloides

Abstract Background The oleaginous yeast Rhodotorula toruloides is a promising chassis organism for the biomanufacturing of value-added bioproducts. It can accumulate lipids at a high fraction of biomass. However, metabolic engineering efforts in this organism have progressed at a slower pace than those in more extensively studied yeasts. Few studies have investigated the lipid accumulation phenotype exhibited by R. toruloides under nitrogen limitation conditions. Consequently, there have been only a few studies exploiting the lipid metabolism for higher product titers. Results We performed a multi-omic investigation of the lipid accumulation phenotype under nitrogen limitation. Specifically, we performed comparative transcriptomic and lipidomic analysis of the oleaginous yeast under nitrogen-sufficient and nitrogen deficient conditions. Clustering analysis of transcriptomic data was used to identify the growth phase where nitrogen-deficient cultures diverged from the baseline conditions. Independently, lipidomic data was used to identify that lipid fractions shifted from mostly phospholipids to mostly storage lipids under the nitrogen-deficient phenotype. Through an integrative lens of transcriptomic and lipidomic analysis, we discovered that R. toruloides undergoes lipid remodeling during nitrogen limitation, wherein the pool of phospholipids gets remodeled to mostly storage lipids. We identify specific mRNAs and pathways that are strongly correlated with an increase in lipid levels, thus identifying putative targets for engineering greater lipid accumulation in R. toruloides . One surprising pathway identified was related to inositol phosphate metabolism, suggesting further inquiry into its role in lipid accumulation. Conclusions Integrative analysis identified the specific biosynthetic pathways that are differentially regulated during lipid remodeling. This insight into the mechanisms of lipid accumulation can lead to the success of future metabolic engineering strategies for overproduction of oleochemicals.

59 BASIC BIOLOGICAL SCIENCES↗

Genome_shuffling_enables_quantitative_trait_locus_mapping_in_Bacillus_subtilis

Genetic mapping is a powerful tool for eukaryotic genetics that has only been applied to bacteria in limited circumstances. Quantitative trait locus (QTL) mapping generally relies on sexual recombination to break linkages between genes, yet bacteria rarely undergo sufficient homologous recombination to generate suitable mapping populations. In this work, we used iterative biparental genome shuffling by protoplast fusion inBacillus subtilisto generate a population of bacteria with substantial random recombination throughout their genomes. Individual shuffled progeny were arrayed in well plates, resequenced, and characterized for a range of complex phenotypes including spore germination and swarming motility. Genetic mapping of the resulting phenotypes identified high-confidence QTLs of moderate size (∼10 kb), and these associations were validated through targeted genetic swaps. ThisB. subtilisQTL population can easily be used to map additional phenotypes, and the general approach for QTL mapping is applicable in a wide range of bacteria.

Bacillus subtilis↗

Optimizing resource allocation in Miscanthus breeding via sparse testing designs for genomic prediction

Phenotyping high-biomass perennial crops is laborious and the rate of genetic gain in conventional perennial crop breeding programs is typically low. So, it is especially important to identify methods that produce efficiency gains in the breeding process. Miscanthus is a C4 perennial grass with favorable characteristics for producing biomass as a feedstock for biofuels and diverse bio-based products. Increasing biomass yield will increase profitability and environmental benefits, so it is a key target for Miscanthus breeding. In addition, the identification of well-adapted genotypes across a wide range of environmental conditions requires the establishment of multi-environment trials (METs). Sparse testing is a genomic prediction-based strategy that reduces the phenotyping costs in METs by selecting a subset of genotypes to evaluate in a subset of environments and then predicts the performance of the unobserved genotype-environment combinations. A Miscanthus sacchariflorus (MSA) population comprising 336 genotypes observed across three environments was analyzed implementing sparse testing designs. Three prediction models considering main effects (environments, genotypes, genomic) and interaction effects (genotype-by-environment; G×E interaction) were implemented for forecasting dry biomass yield (YDY), total culm (TCM), average internode length (AIL), and culm node number (CNN). Multiple calibration sets based on different compositions and sizes were considered to evaluate performance in terms of the predictive ability (PA) and the mean square error (MSE) for a fixed testing set size. The training set size ranged from 52 to 112 to predict a fixed set of 224 unobserved genotypes across all three environments. The results showed that the model accounting for G×E interaction consistently presented the highest PA and the lowest MSE: for CNN (PA: ~0.77, MSE: ~0.5) and YDY (PA: ~0.70, MSE: ~1.3) while for TCM and AIL these ranged from ~0.28 to 0.41 and ~1.3 to 4.3, respectively. Overall, varying training sets and allocation strategies did not affect PA and MSE, with 52 non-overlapping and 0 overlapping genotypes per environment as the optimal cost-effective allocation framework. This suggests that implementing sparse testing designs could significantly reduce phenotyping costs by fivefold, without compromising PA in breeding programs for perennial crops such as Miscanthus.

Miscanthus sacchariflorus (MSA)↗

Convergent evolution of aerobic fermentation through divergent mechanisms acting on the same key glycolytic genes

Expression profiling by high throughput sequencing The model yeast Saccharomyces cerevisiae has evolved aerobic fermentation in part through a whole genome duplication and subsequent retention of glycolytic genes. To identify independent yeast lineages with a high glycolytic rate phenotype we developed an assay measuring extracellular acidification rates (ECAR) to evaluate the glycolytic rates across diverse yeast species. This assay identified a novel group of yeasts in the genus Saturnispora with rapid glycolytic rates. Through comparative approaches, we found that several glycolytic genes encoding hexokinases and enzymes in lower glycolysis had higher expression and modifications in promoter sequences in rapid ECAR species. Intriguingly, many of the upregulated genes are the same genes that are duplicated in S. cerevisiae. When the transcription factor required for their activation was deleted, the mutants had a slow glycolytic rate and more respiratory phenotype. This work shows how the aerobic fermentation phenotype has convergently evolved in Saturnispora through transcriptional rewiring. This divergent mechanism nevertheless impacts the same glycolytic genes, which suggests that there are evolutionary constraints on how aerobic fermentation can arise.

Horianopoulos, Linda C. [Department of Food Scienc↗