Biology subjects

Scholes, A. N.

Publications and source records attributed to Scholes, A. N..

6 recordsLinked to original sources

bioRxiv2025

Surprising regulatory plasticity for the conserved HOG pathway in diverse Saccharomyces cerevisiae strains

Mitogen-activated protein kinases (MAPKs) display remarkable regulatory plasticity across evolution, ranging from highly specialized pathways to broadly responsive global signaling hubs. In the budding yeast Saccharomyces cerevisiae, the high-osmolarity glycerol (HOG) network has served as a paradigm for largely stress-specific MAPK signaling, where the Hog1 MAPK coordinates osmoadaptation. This stands in sharp contrast to Hog1 orthologs in other fungi and humans, which respond not only to osmotic stress, but also diverse stresses including UV, heat shock, oxidative stress, and pathogen signals. Whether the relative osmospecificity of S. cerevisiae Hog1 represents an ancestral feature or lineage-specific evolution remains unclear. The majority of foundational work on S. cerevisiae HOG signaling has been performed in laboratory strains that are known to be genetic and phenotypic outliers, and we have been leveraging wild S. cerevisiae strains to understand aspects of stress signaling that may have been lost in laboratory strains. Here, we examined the phenotypic effects of hog1{Delta} mutations in a commonly-used laboratory S. cerevisiae strain and diverse wild strains on stress cross protection and gene expression. Our findings demonstrate an expanded role in cross-stress protection for Hog1 in wild yeast strains compared to the laboratory strain. More strikingly, we identified a large number of Hog1-dependent genes for non-osmotic stresses in the wild strains that were completely absent in the lab strain. Notably, the Hog1 regulon in wild strains responding to non-osmotic stresses is largely distinct from the canonical osmotic stress response, which we show likely occurs through non-canonical cytoplasmic functions. These findings reveal surprising within-species plasticity for the highly conserved HOG network, suggesting that evolutionary transitions between specialist to generalist stress signaling may occur with relative ease.

genetics↗

bioRxiv2023

Natural variation in yeast reveals multiple paths for acquiring higher stress resistance

BackgroundOrganisms frequently experience environmental stresses that occur in predictable patterns and combinations. For wild Saccharomyces cerevisiae yeast growing in natural environments, cells may experience high osmotic stress when they first enter broken fruit, followed by high ethanol levels during fermentation, and then finally high levels of oxidative stress resulting from respiration of ethanol. Yeast have adapted to these patterns by evolving sophisticated "cross protection" mechanisms, where mild primary doses of one stress can enhance tolerance to severe doses of a different secondary stress. For example, in many yeast strains, mild osmotic or mild ethanol stresses cross protect against severe oxidative stress, which likely reflects an anticipatory response important for high fitness in nature. ResultsDuring the course of genetic mapping studies aimed at understanding the mechanisms underlying natural variation in ethanol-induced cross protection against H2O2, we found that a key H2O2 scavenging enzyme, cytosolic catalase T (Ctt1p), was absolutely essential for cross protection in a wild oak strain. This suggested the absence of other compensatory mechanisms for acquiring H2O2 resistance in that strain background under those conditions. In this study, we found surprising heterogeneity across diverse yeast strains in whether CTT1 function was fully necessary for acquired H2O2 resistance. Some strains exhibited partial dispensability of CTT1 when ethanol and/or salt were used as mild stressors, suggesting that compensatory peroxidases may play a role in acquired stress resistance in certain genetic backgrounds. We leveraged global transcriptional responses to ethanol and salt stresses in strains with different levels of CTT1 dispensability, allowing us to identify possible regulators of these alternative peroxidases and acquired stress resistance in general. ConclusionsUltimately, this study highlights how superficially similar traits can have different underlying molecular foundations and provides a framework for understanding the diversity and regulation of stress defense mechanisms. Author SummaryOrganisms in nature frequently experience environmental stress in predictable patterns. For example, during the summer months in temperate climates, the warmth of the morning sun gradually gives way to high afternoon temperatures. Organisms that can anticipate these predictable patterns to mobilize stress defenses would have an advantage in nature. One way organisms anticipate future stress is through cross protection, where cells exposed to a mild dose of one stress gain the ability to survive an otherwise lethal dose of different stress. To better understand the molecular mechanisms that are responsible cross protection, we have been taking advantage of wild yeast strains that are either more resilient or more sensitive to stresses. During the course of this study, we found that strains with superficially similar levels of cross protection differ in the precise molecular mechanisms that underlie the trait. Our study suggests that different molecular strategies may be important for yielding similar stress resistances under different environmental constraints, and highlights the power of harnessing natural genetic diversity to understand the molecular mechanisms underlying differences in environmental responses.

genomics↗

bioRxiv2023

A Hypothesis-Based Hop Microbiology Laboratory Module Testing the Plausibility of the Mythical Origin of the India Pale Ale (IPA)

As one of the most famous fermented drinks in the world, beer is an especially relatable topic for microbiology courses. Here, we describe a short and easily adaptable module based on the antibacterial properties of hops used in brewing. By the 15th century, beer recipes included hops (the flower of the Humulus lupulus plant) as a bittering agent and antimicrobial. By the 19th century, the highly-hopped Indian Pale Ale (IPA) became popular, and a modern myth has emerged that IPAs were invented to survive long ocean voyages such as from Britain to India. With that myth in mind, we designed a hypothesis-driven microbiology lab module that tests the plausibility of this brewing myth-- namely that highly-hopped beers possess enough antibacterial activity to prevent spoilage, while lowly-hopped beers do not. The overall design of the module is to test the antimicrobial properties of hops using petri plates containing varying concentrations of hop extract. The module includes hypothesis generation and testing related to bacterial physiology and morphology (hops are not equally effective against gram-positive and gram-negative bacteria), and to mechanisms of antimicrobial resistance (as beer spoilage bacteria have repeatedly evolved hop resistance). Pre and post assessment showed that students made significant gains in the learning objectives for the module, which encourages critical thinking and hypothesis testing by linking microbial physiology and antimicrobial resistance to an important and topical real-world application.

scientific communication and education↗

bioRxiv2019

Comparison of RNA Isolation Methods on RNA-Seq: Implications for Differential Expression and Meta-Analyses

Technical variation across different batches of RNA-seq experiments can clearly produce spurious signals of differential expression and reduce our power to detect true differences. Thus, it is important to identify major sources of these so-called \"batch effects\" to eliminate them from study design. Based on the different chemistries of \"classic\" phenol extraction of RNA compared to common commercial RNA isolation kits, we hypothesized that specific mRNAs may be preferentially extracted depending upon method, which could masquerade as differential expression in downstream RNA-seq analyses. We tested this hypothesis and found that phenol extraction preferentially isolated membrane-associated mRNAs, thus resulting in spurious signals of differential expression. Within a self-contained experimental batch (e.g. control versus treatment), the method of RNA isolation had little effect on the ability to identify differentially expressed transcripts. However, we suggest that researchers performing meta-analyses across different experimental batches strongly consider the RNA isolation methods for each experiment.

genomics↗

bioRxiv2019

A Wild Yeast Laboratory Activity: From Isolation to Brewing

Microbial fermentation is a common form of metabolism that has been exploited by humans to great benefit. Industrial fermentation currently produces a myriad of products ranging from biofuels to pharmaceuticals. About one third of the worlds food is fermented, and the brewing of fermented beverages in particular has an ancient and storied history. Because fermentation is so intertwined with our daily lives, the topic is easily relatable to students interested in real-world applications for microbiology. Here, we describe the curriculum for an inquiry-based laboratory course that combines yeast molecular ecology and brewing. The rationale for the course is to compare commercial Saccharomyces cerevisiae yeast strains, which have been domesticated through thousands of generations of selection, with wild yeast, where there is growing interest in their potentially unique brewing characteristics. Because wild yeast are so easy to isolate, identify, and characterize, this is a great opportunity to present key concepts in molecular ecology and genetics in a way that is relevant and accessible to students. We organized the course around three main modules: isolation and identification of wild yeast, phenotypic characterization of wild and commercial ale yeast strains, and scientific design of a brewing recipe and head-to-head comparison of the performance of a commercial and wild yeast strain in the brewing process. Pre and post assessment showed that students made significant gains in the learning objectives for the course, and students enjoyed connecting microbiology to a real-world application.

scientific communication and education↗

bioRxiv2017

Linkage mapping of yeast cross protection connects gene expression variation to a higher-order organismal trait

Gene expression variation is extensive in nature, and is hypothesized to play a major role in shaping phenotypic diversity. However, connecting differences in gene expression across individuals to higher-order organismal traits is not trivial. In many cases, gene expression variation may be evolutionarily neutral, and in other cases expression variation may only affect phenotype under specific conditions. To understand connections between gene expression variation and stress defense phenotypes, we have been leveraging extensive natural variation in the gene expression response to acute ethanol in laboratory and wild Saccharomyces cerevisiae strains. Previous work found that the genetic architecture underlying these expression differences included dozens of \"hotspot\" loci that affected many transcripts in trans. In the present study, we provide new evidence that one of these expression QTL hotspot loci is responsible for natural variation in one particular stress defense phenotype--ethanol-induced cross protection against severe doses of H2O2. The causative polymorphism is in the heme-activated transcription factor Hap1p, which we show directly impacts cross protection, but not the basal H2O2 resistance of unstressed cells. This provides further support that distinct cellular mechanisms underlie basal and acquired stress resistance. We also show that the Hap1p-dependent cross protection relies on novel regulation of cytosolic catalase T (Ctt1p) during ethanol stress in wild strains. Because ethanol accumulation precedes aerobic respiration and accompanying reactive oxygen species formation, wild strains with the ability to anticipate impending oxidative stress would likely be at an advantage. This study highlights how strategically chosen traits that better correlate with gene expression changes can improve our power to identify novel connections between gene expression variation and higher-order organismal phenotypes.

genetics↗