Biology subjects

Ridgway, M.

Publications and source records attributed to Ridgway, M..

6 recordsLinked to original sources

bioRxiv2026

Genetic variation in behavioral and physiological responses to copper in Drosophila melanogaster

Anthropogenic copper (Cu) contamination from agriculture, mining, and industrial runoff creates environmental gradients affecting physiology and behavior in wild populations. While Cu toxicity in Drosophila melanogaster is well characterized, it remains unclear whether Cu resistance is one integrated trait or several independently evolving components. Using a subset of recombinant inbred lines (RILs) from the Drosophila Synthetic Population Resource (DSPR), we measured three components of Cu response: feeding avoidance, oviposition avoidance, and physiological tolerance (median lethal time, LT50) under sustained Cu exposure. All three traits showed substantial phenotypic variation among RILs. Feeding and oviposition avoidance were both highly heritable (H 2 ~ 0.88), and RIL identity accounted for 49.5% of the variance in LT50. However, the three traits showed no significant correlation across RILs, indicating distinct genetic architecture. We identified a single male specific quantitative trait locus (QTL) on chromosome 2R that explained 17.7% of the variation in feeding preference; the interval included candidate detoxification genes Jheh1, Jheh2, Jheh3 and sano, the latter of which is associated with olfactory behavior. No significant QTL were detected for oviposition preference, suggesting a highly polygenic structure that may difficult to detect with our limited panel size. Together, these results indicate that Cu resistance in D. melanogaster is genetically modular. Behavioral avoidance during feeding, oviposition, and physiological tolerance are heritable but architecturally distinct components, each with potential to respond to selection independently.

genetics↗

bioRxiv2025

Origins and consequences of kinetoplast loss in trypanosomes

The kinetoplast is the large mitochondrial genome present in the eponymous Kinetoplastida. Trypanosoma brucei is an African trypanosome that can lose kinetoplast DNA (kDNA), however, when the nuclear-encoded gamma subunit of the mitochondrial F1FO-ATP synthase ({gamma}ATPase) is mutated. These mutations, analogous to a broken camshaft at the core of the ATP synthase rotary motor, are associated with multidrug resistance, and correlated with tsetse-fly independent mechanical transmission, and geographical spread of these parasites beyond Africa. Here we engineer kinetoplast-independent T. brucei to explore origins and consequences of kDNA loss. We used oligo targeting to edit the native{gamma} ATPase gene, and selection with the ATP synthase targeting drug oligomycin to enrich the desired mutants. Using this approach, we identified novel M282F, M282W, and M282Y mutants, and subsequently generated precision-edited strains expressing the previously described L262P or A273P mutants, or the novel M282F mutant. Heterozygous M282F mutants retained sensitivity to the kDNA-targeting drug acriflavine, while homozygous M282F mutants were acriflavine resistant and readily tolerated acriflavine-induced kDNA loss. Proteomics analysis of the homozygous mutant pre-kDNA-loss revealed highly specific depletion of ATP synthase-associated proteins, but not the F1 subunits. Complete kDNA-loss in these cells was associated with substantial depletion of kDNA-binding proteins and mitochondrial RNA-processing factors. In contrast, mitochondrial membrane-associated transporters were increased in abundance. We conclude that T. brucei cells with a homozygous{gamma} ATPase M282F mutation assemble a remodelled ATP synthase and readily tolerate kDNA loss, which is accompanied by substantial remodelling of the mitochondrial proteome Author summaryMutations in the gamma subunit of the mitochondrial ATP synthase in parasitic African trypanosomes can have major consequences. Specifically, the entire large and complex mitochondrial genome, the kinetoplast, is rendered dispensable, and the cells become resistant to important kinetoplast-targeting drugs. Veterinary parasites with these mutations have also spread outside Africa through simple mechanical transmission, either sexually or by biting flies or vampire bats. We precision-edited the gamma subunit to replicate previously described mutants and identified a novel mutant that readily tolerated kinetoplast loss. Using quantitative proteomics, we demonstrated highly specific depletion of ATP synthase-associated proteins pre-kinetoplast-loss. We then use genome sequencing to show that the kinetoplast could be completely lost by these cells and demonstrated that cells lacking mitochondrial nucleic acids displayed specific depletion of mitochondrial nucleic acid-binding proteins. Notably, several mitochondrial membrane-associated transporter complexes were increased in abundance. Thus, we establish a method to test precise {gamma}ATPase mutations and to identify new mutations associated with kinetoplast loss. We also show that trypanosomes with a dispensable kinetoplast specifically remodel the ATP synthase pre-kinetoplast-loss and substantially remodel the mitochondrial proteome post-kinetoplast-loss.

microbiology↗

bioRxiv2025

Acoziborole resistance associated mutations in trypanosome CPSF3

Acoziborole is a safe, single dose, oral therapy, for treatment of both early and late-stage sleeping sickness, a deadly disease caused by African trypanosomes. Other benzoxaboroles show efficacy against other trypanosomatids, apicomplexans, fungi, bacteria, and viruses. Acoziborole targets the trypanosome pre-mRNA processing endonuclease, cleavage and polyadenylation specificity factor 3 (CPSF3), and triggers CPSF3 degradation, but it remains unclear whether additional mechanisms contribute to efficacy. We used oligo targeting for site saturation mutagenesis of the native CPSF3 gene. Among >1,500 edits around the putative drug binding site, only Asn232His edits conferred moderate resistance to acoziborole. Using a novel combinatorial oligo targeting method we edited multiple sites simultaneously, including sites that differ in human CPSF3, and found that an Asn232His, Tyr383Phe, Asn448Gln triple-mutant strain was >40-fold resistant to acoziborole. We used gene tagging to show that all three edits were on the same allele, and to show that triple-mutant CPSF3 was highly resistant to rapid acoziborole and proteasome-dependent degradation. Computational modelling revealed how the combinatorial mutations can disrupt acoziborole - CPSF3 interactions by introducing steric clash and by disrupting hydrophobic and water-mediated interactions. We conclude that acoziborole safety and efficacy can be explained by selective affinity for, and rapid turnover of, trypanosome CPSF3. Author SummaryDiagnosis and treatment options, previously limited for sleeping sickness, have been transformed in recent years. Acoziborole, for example, is a new, safe, single dose, oral therapy for the treatment of this deadly disease. This drug can also be used without the need for cumbersome disease-stage diagnosis. Additional boron-based drugs also show great promise against a whole range of other infectious diseases. Acoziborole targets an RNA processing enzyme in African trypanosomes, and triggers its degradation, but human cells express a similar enzyme, and alternative trypanosomal targets have also been suggested. Insights into how a drug interacts with its target can help to understand selective action against a pathogen, and to predict resistance, an ever-present threat for many drugs. We used a precision gene editing method to change the target protein in trypanosomes, editing single sites or multiple sites simultaneously. A novel triple-mutant was found to be both highly resistant to acoziborole and highly resistant to rapid degradation. Using computational models, we were able to explain how multiple mutations interfered with acoziborole binding to its target. The findings show how selective binding of a specific parasite enzyme makes acoziborole such a safe and effective drug.

microbiology↗

bioRxiv2025

Fish spawning events stimulate trophic hotspots across freshwater food webs

1.0Mass reproductive events, where large numbers of organisms aggregate for synchronous reproduction, are often captivating and ecologically significant, boosting offspring survival and reducing time spent searching for mates. However, mass reproduction can also instigate cryptic but consequential responses across entire food webs. Reproductive materials are abundant and accessible resources that can attract mobile consumers from up to thousands of kilometers away. Yet, their consumption, especially in aquatic systems, is difficult to detect and rarely characterized. Here, we combine molecular techniques with acoustic telemetry, literature review, and extensive natural history observations to investigate the food web consequences of synchronized reproduction in freshwater fishes. First, we demonstrate that a common but underappreciated fish species, white sucker, creates a resource pulse used ubiquitously by consumers, from local invertebrates and fishes to mobile predatory fishes, birds, and terrestrial mammals. Spawning white sucker create trophic hotspots that attract consumers across trophic levels and ecosystems to feed on eggs, spawning adults, and aggregated egg predators. Then we show that egg provisioning and predation is widespread among north-temperate freshwater fish species, highlighting that resource pulses instigated by mass reproduction may play a critical but underappreciated role in freshwater and terrestrial ecosystems.

ecology↗

bioRxiv2025

Decoding efficacy and resistance space at a drug binding site

Interactions between drugs and their targets impact efficacy and, when altered by mutation, can result in resistance 1-3. Assessing and understanding the impacts of all possible mutations at a drug binding site remain challenging, however 4-6. Here, we used Multiplex Oligo Targeting (MOT) for mutational profiling, and computational modelling, to decode efficacy and resistance space at the otherwise native binding site for a low nanomolar potency, anti-trypanosomal, proteasome inhibitor 7. We saturation-edited twenty codons in the Trypanosoma brucei proteasome {beta}5 subunit and subjected the resulting MOT libraries to stepwise drug selection. Amplicon sequencing, and codon variant scoring, yielding dose-response profiles for >100 resistance-conferring mutants, among 1,280 possible codon variants. Codon variant scores were predictive of relative resistance observed using a bespoke set of mutants, while fitness profiling revealed otherwise extensive constraints on mutational fitness and resistance space. The resistance profile that emerged allowed us to readily predict routes to spontaneous drug resistance observed within accessible, single nucleotide mutational space. In silico analysis of {beta}5 subunit mutations predicted impacts on ligand affinity via steric effects, hydrogen-bonding and lipophilicity, which when combined with predictions of proteasome function perturbing mutations, were closely aligned with observed impacts on drug resistance. We conclude that MOT-library profiling facilitates assessment of all possible mutations at a drug binding site. Further decoding of drug target structure-activity relationships and drug resistance space will facilitate the design of more effective and durable drugs.

molecular biology↗

bioRxiv2019

Winter in water: Differential responses and the maintenance of biodiversity

The ecological consequences of winter in freshwater systems are an understudied but rapidly emerging research area. Here, we argue that winter periods of reduced temperature and light (and potentially oxygen and resources) could play an underappreciated role in mediating the coexistence of species. This may be especially true for temperate and subarctic lakes, where seasonal changes in the thermal environment might fundamentally structure species interactions. With climate change already shortening ice-covered periods on temperate and polar lakes, consideration of how winter conditions shape biotic interactions is urgently needed. Using freshwater fishes in northern temperate lakes as a case study, we demonstrate how physiological trait differences (e.g., thermal preference, light sensitivity) drive differential behavioral responses to winter among competing species. Specifically, some species have a higher capacity for winter activity than others. Existing and new theory is presented to argue that such differential responses to winter can promote species coexistence. Importantly, if winter is a driver of niche differences that weaken competition between relative to within species, then shrinking winter periods could threaten coexistence by tipping the scales in favor of certain sets of species over others.

ecology↗