Traumatic brain injury is a major cause of chronic neurological impairment worldwide, and there is evidence that both genetic and environmental variation contribute to the likelihood of recovery. Using an insect model of traumatic brain injury, we examined variation in the risk of mortality using quantitative genetic approaches applied previously for life history traits in Drosophila melanogaster. We quantified additive genetic variance for mortality risk using a controlled breeding design and found levels of variation consistent with existing data on major fitness components. We did not detect inbreeding depression for mortality risk, suggesting that this trait is not strongly affected by recessive deleterious alleles. To explain the high level of standing genetic variation, we considered whether mortality risk depends on the metabolic resources available to an individual, also known as “condition”. We manipulated condition by inducing random mutations and by restricting calories during larval development. We found that reduced condition due to both random mutations and resource limitation significantly increased the risk of mortality following trauma. Among inbred lines, greater mortality risk was associated with lower viability, fecundity and longevity, consistent with an effect of genome-wide genetic quality. Our results suggest that further consideration of individual condition would be valuable for understanding and predicting variation in the outcomes of traumatic brain injury.

The impact of selection versus genetic drift on the evolution of mutation patterns is unclear. In Saccharomyces cerevisiae, which is predominantly diploid in nature, there is evidence that haploid cells have a higher mutation rate than diploids, suggesting that a haploid-specific mutator phenotype may have evolved due to the limited opportunity for selection to act on this rare cell type. Mutation in haploids was primarily elevated in late-replicating regions of the genome, implicating error-prone translesion synthesis (TLS) repair. Additional research has demonstrated that removing REV1, a gene responsible for initiating TLS, causes a reduction in haploid mutation rate. To assess whether the preferential use of this error-prone repair pathway by haploids explains the difference in genome-wide mutation patterns between cell types, we deleted REV1 in both diploid and haploid S. cerevisiae and estimated their mutation rates using a mutation accumulation experiment. Consistent with a previous study, we found a 50% higher single nucleotide mutation rate in REV1+ haploids than in REV1+ diploids. Deleting the REV1 gene caused this difference to vanish, with mutation rates in haploid and diploid rev1Δ lines converging on 2.4 × 10-10. Our results suggest that the mutagenic effect of translesion synthesis is much stronger in haploids, reflecting a limited opportunity for selection to act on mutation rates in rarer cells or smaller populations. We also find evidence that REV1 plays an important role in mitochondrial genome maintenance in both cell types.

Traumatic brain injury (TBI) is a leading cause of disability and death, with outcome severity varying widely even among individuals with comparable injuries. A major challenge is to identify pathways that underlie this variation and could be targeted to improve therapies. Innate immune pathways are candidates because they are rapidly activated after TBI and contribute to neurodegenerative disorders. Using a Drosophila melanogaster TBI model, we examined how genetic background, age, and diet modify effects of evolutionarily conserved Toll and Immune deficiency (Imd) pathways on injury outcomes. These pathways signal through nuclear factor-kappa B (NF-κB) transcription factors Dorsal-related immunity factor (Dif) and Relish (Rel) to activate antimicrobial peptide (AMP) gene expression. We found that genetic diversity among lines from the Drosophila Genetic Reference Panel (DGRP) contributed to variation in AMP expression before and after TBI, with additional effects of age and diet. AMP expression tended to be correlated positively with early mortality following TBI in young flies, but negatively in older flies, suggesting an age-dependent shift in AMP effects from detrimental to protective. Furthermore, heterozygous mutations in Dif or Rel lowered AMP expression in a diet-dependent manner and led to correspondingly reduced early mortality after TBI. These findings show that genetic, biological, and environmental factors influence innate immune pathways, which in turn determine TBI outcomes. Innate immune gene expression before injury emerges as a potential prognostic indicator, pointing to potential new therapeutic strategies.

There is significant variation in the rate and spectrum of spontaneous mutations among taxa. How this variation is shaped by natural selection remains a subject of debate. The drift barrier hypothesis proposes that selection generally favors lower mutation rates due to the risk of deleterious mutations but acts less effectively against weak mutator alleles in smaller populations, allowing the mutation rate to increase due to genetic drift. Given this model, we propose that mutation rates may also be elevated in cell types that appear rarely in a population, where DNA replication and repair processes are subject to selection less often. We can begin to test this prediction in yeast species, some of which can be grown in either a haploid or diploid cell state. Existing data on the budding yeast Saccharomyces cerevisiae support this prediction, with a higher mutation rate observed in haploids, which is the rare cell type in natural populations. However, this pattern could also appear if haploidy is inherently mutagenic, regardless of the dominant cell type. To test these alternatives, we conducted a mutation accumulation experiment with haploid and diploid cells of the fission yeast Schizosaccharomyces pombe, in which diploidy is the rare cell type. In this species, we found a higher mutation rate in diploids, consistent with our prediction. In both species, the spectrum of mutations is also influenced by ploidy state. Our findings suggest that limits to selection on mutation may be evident as variation within species.

The impact of selection versus genetic drift on the evolution of mutation patterns is unclear. In Saccharomyces cerevisiae , which is predominantly diploid in nature, there is evidence that haploid cells have a higher mutation rate than diploids, suggesting that a haploid-specific mutator phenotype may have evolved due to the limited opportunity for selection to act on this rare cell type. Mutation in haploids was primarily elevated in late-replicating regions of the genome, implicating error-prone translesion synthesis (TLS) repair. Additional research has demonstrated that removing REV1 , a gene responsible for initiating TLS, causes a reduction in haploid mutation rate. To assess whether the preferential use of this error-prone repair pathway by haploids explains the difference in genome-wide mutation patterns between cell types, we deleted REV1 in both diploid and haploid S. cerevisiae and estimated their mutation rates using a mutation accumulation experiment. Consistent with a previous study, we found a 50% higher single nucleotide mutation rate in REV1 + haploids than in REV1 + diploids. Deleting the REV1 gene caused this difference to vanish, with mutation rates in haploid and diploid rev1 Δ lines converging on 2.4 × 10 -10 . Our results suggest that the mutagenic effect of translesion synthesis is much stronger in haploids, reflecting a limited opportunity for selection to act on mutation rates in rarer cells or smaller populations. We also find evidence that REV1 plays an important role in mitochondrial genome maintenance in both cell types.

Sexual selection contributes to biodiversity and the costs and benefits of sexual reproduction. In organisms where sex is infrequent, these impacts of sexual selection are likely to be limited. An increased frequency of obligate sex would increase the opportunity for sexual selection, which could promote the evolution of sexual traits and sexual differentiation. To study these dynamics, we conducted experimental evolution in the yeast Saccharomyces cerevisiae, which is predominantly asexual, with two isogamous mating types. We used selectable markers to impose frequent obligate sex in 96 populations. We manipulated the opportunity for sexual selection by imposing skewed mating-type ratios, either enforcing an alternation of haploid and diploid growth or allowing unrestricted mating following sporulation. After just ten sexual cycles, we observed evolution in growth, cell size, pheromone production, and mating, with the mating types responding asymmetrically, but little evolutionary change in sporulation rate. Mating type dimorphism increased, with evident trade-offs between growth, attractiveness, and cell size. Genome sequences from a subset of populations revealed many mutations affecting sex-related genes. Unexpectedly, when alternation of ploidy states was not enforced, the populations evolved to become sporulation-competent haploids, unlinking meiosis from ploidy change. Our results illustrate that sexual differentiation can evolve rapidly in response to an increased opportunity for sexual selection.

Genomic regions containing tandem duplications may be subject to particularly high rates of copy number mutations due to recombinational repair of DNA damage. Consequently, changes in copy number may be among the most accessible beneficial mutations during evolution in a new environment. In the budding yeast Saccharomyces cerevisiae, the gene CUP1 occurs in multiple tandem copies and encodes a metallothionein relevant for copper homeostasis. We examined CUP1 copy number in 220 mutation accumulation lines and their ancestors to quantify spontaneous genetic change at this locus. We also measured copper tolerance in these strains to understand the phenotypic effects of CUP1 copy number change and other mutations. Mutations in CUP1 copy number occurred relatively rapidly, with a bias towards losses. While copper tolerance also declined due to mutations, this change was less rapid and was only partly driven by CUP1 copy number. Thus, while CUP1 is highly susceptible to copy number change, our results add to a body of evidence that these accessible mutations on their own do not fully account for variation in copper tolerance.

Of the array of spontaneous mutations that can occur, changes to chromosome number may have the greatest impact on the evolutionary potential of populations and the condition of affected individuals. Chromosomal nondisjunction resulting in aneuploidy is found across eukaryotes, but the consequences of such karyotypic variation have not been widely explored. In the fruit fly Drosophila melanogaster, aneuploid females with an XXY karyotype can arise through nondisjunction, inheriting a Y chromosome from their male parent. While the Y chromosome contains few genes, the large amount of heterochromatic DNA it contains can substantially alter genome-wide gene expression in females. We conducted a series of experiments to understand how sex chromosome aneuploidy alters key traits in affected females and their progeny. In the same genetic background, we also determined the rate at which this karyotype appears spontaneously and its standing frequency. We found that XXY females largely resembled XX females, but experienced size and fecundity benefits when receiving a male-transmitted Y chromosome. However, XYY males produced by aneuploid females experienced reduced viability, limiting the standing frequency of aneuploidy at mutation-selection equilibrium. Our findings demonstrate that aneuploid flies are not too rare in laboratory populations, but that the effects of this karyotypic diversity depend on sex and parent of origin.

Environmental conditions can influence mutation rates, but the reasons are often unclear. Budding yeast can utilize many carbon sources, with variation in the degree of fermentation versus respiration. Since aerobic respiration produces mutagenic reactive oxygen species, we hypothesized that yeast grown in media promoting aerobic respiration would show higher mutation rates. We found significant differences across five media types, with the highest mutation rate in pyruvate and the lowest in glucose. However, mutation rates responded to respiration rate in a nonlinear fashion, suggesting that the degree of respiration in a given environment is only partly predictive of mutation rate.

Aneuploidy, arising from the gain or loss of chromosomes due to nondisjunction, is a special class of mutation. It can create significant phenotypic changes by altering the abundance of hundreds of genes in a single event, providing material for adaptive evolution. But it can also incur large fitness costs relative to other types of mutations. Understanding the mutational dynamics of aneuploidy is important for modeling its impact in nature, but aneuploidy rates are difficult to measure accurately. One challenge is that aneuploid karyotypes may revert back to euploidy, biasing forward mutation rate estimates-yet the rate of aneuploidy reversion is largely uncharacterized. Furthermore, current rate estimates are confounded because fitness differences between euploids and aneuploids are typically not accounted for in rate calculations. We developed a unique fluctuation assay in a wild-yeast model to measure the rate of extra-chromosome loss across 3 aneuploid chromosomes while accounting for fitness differences between aneuploid and euploid cells. We show that incorporating fitness effects is essential to obtain accurate estimates of aneuploidy rates. Furthermore, the rate of extra-chromosome loss, separate from karyotype fitness differences, varies across chromosomes. We also measured rates in a strain lacking RNA-binding protein Ssd1, important for aneuploidy tolerance and implicated in chromosome segregation. We found no role for Ssd1 in the loss of native aneuploid chromosomes, although it did impact an engineered chromosome XV with a perturbed centromeric sequence. We discuss the impacts and challenges of modeling aneuploidy dynamics in real-world situations.

In populations with separate sexes, genetic load due to deleterious mutations may be expressed differently in males and females. Evidence from insect models suggests that selection against mutations is stronger in males. This pattern will reduce deleterious allele frequencies at the expense of males, such that female mean fitness is greater than expected, preserving population persistence in the face of high mutation rates. While previous studies focus on reproductive success, mutation load depends on total selection in each sex, including selection for viability. We might expect minimal sex differences in viability effects in fruit flies, since male and female larvae behave similarly, yet many genes show sex-biased expression in larvae. We measured the sex-specific viability effects of nine "marker" mutations and 123 mutagenized chromosomes. We find that both types of mutations generally reduce viability in both sexes. Among marker mutations we detect instances of sex-biased effects in each direction; mutagenized chromosomes show little sex-specific mutational variance, but recessive lethals show a female bias, including in FlyBase records. We conclude that mutations regularly affect viability in a sex-specific manner, but that the strong pattern of male-biased mutational effects observed previously for reproductive success is not apparent at the pre-reproductive stage.

The role of spontaneous mutations in evolution depends on the distribution of their effects on fitness. Despite a general consensus that new mutations are deleterious on average, a handful of mutation accumulation experiments in diverse organisms instead suggest that beneficial and deleterious mutations can have comparable fitness impacts, i.e. the product of their respective rates and effects can be roughly equal. We currently lack a general framework for predicting when such a pattern will occur. One idea is that beneficial mutations will be more evident in genotypes that are not well adapted to the testing environment. We tested this prediction experimentally in the laboratory yeast Saccharomyces cerevisiae by allowing nine replicate populations to adapt to novel environments with complex sets of stressors. After >1000 asexual generations interspersed with 41 rounds of sexual reproduction, we assessed the mean effect of induced mutations on yeast growth in both the environment to which they had been adapting and the alternative novel environment. The mutations were deleterious on average, with the severity depending on the testing environment. However, we found no evidence that the adaptive match between genotype and environment is predictive of mutational fitness effects.

The ribosomal DNA array in Saccharomyces cerevisiae consists of many tandem repeats whose copy number is believed to be functionally important but highly labile. Regulatory mechanisms have evolved to maintain copy number by directed mutation, but how spontaneous variation at this locus is generated and selected has not been well characterized. We applied a mutation accumulation approach to quantify the impacts of mutation and selection on this unique genomic feature across hundreds of mutant strains. We find that mutational variance for this trait is relatively high, and that unselected mutations elsewhere in the genome can disrupt copy number maintenance. In consequence, copy number generally declines gradually, consistent with a previously proposed model of rDNA maintenance where a downward mutational bias is normally compensated by mechanisms that increase copy number when it is low. This pattern holds across ploidy levels and strains in the standard lab environment but differs under some stressful conditions. We identify several alleles, gene categories, and genomic features that likely affect copy number, including aneuploidy for chromosome XII. Copy number change is associated with reduced growth in diploids, consistent with stabilizing selection. Levels of standing variation in copy number are well predicted by a balance between mutation and stabilizing selection, suggesting this trait is not subject to strong diversifying selection in the wild. The rate and spectrum of point mutations within the rDNA locus itself are distinct from the rest of the genome and predictive of polymorphism locations. Our findings help differentiate the roles of mutation and selection and indicate that spontaneous mutation patterns shape several aspects of ribosomal DNA evolution.

Systematic editing of yeast genes to generate thousands of mutations indicates that, overall, the mutations have similar effects on yeast fitness regardless of whether they change the protein sequences encoded by the mutated genes.

Mutation is the origin of all genetic variation, good and bad. The mutation process can evolve in response to mutations, positive or negative selection, and genetic drift, but how these forces contribute to mutation-rate variation is an unsolved problem at the heart of genetics research. Mutations can be challenging to measure, but genome sequencing and other tools have allowed for the collection of larger and more detailed datasets, particularly in the yeast-model system. We review key hypotheses for the evolution of mutation rates and describe recent advances in understanding variation in mutational properties within and among yeast species. The multidimensional spectrum of mutations is increasingly recognized as holding valuable clues about how this important process evolves.

All adaptive alleles in existence today began as mutations, but a common view in ecology, evolution, and genetics is that non-neutral mutations are much more likely to be deleterious than beneficial and will be removed by purifying selection. By dramatically limiting the effectiveness of selection in experimental mutation accumulation lines, multiple studies have shown that new mutations cause a detectable reduction in mean fitness. However, a number of exceptions to this pattern have now been observed in multiple species, including in highly replicated, intensive analyses. We briefly review these cases and discuss possible explanations for the inconsistent fitness outcomes of mutation accumulation experiments. We propose that variation in the outcomes of these studies is of interest and understanding the underlying causes of these diverse results will help shed light on fundamental questions about the evolutionary role of mutations.

Predicting fitness in natural populations is a major challenge in biology. It may be possible to leverage fast-accumulating genomic data sets to infer the fitness effects of mutant alleles, allowing evolutionary questions to be addressed in any organism. In this paper, we investigate the utility of one such tool, called PROVEAN. This program compares a query sequence with existing data to provide an alignment-based score for any protein variant, with scores categorized as neutral or deleterious based on a pre-set threshold. PROVEAN has been used widely in evolutionary studies, for example, to estimate mutation load in natural populations, but has not been formally tested as a predictor of aggregate mutational effects on fitness. Using three large published data sets on the genome sequences of laboratory mutation accumulation lines, we assessed how well PROVEAN predicted the actual fitness patterns observed, relative to other metrics. In most cases, we find that a simple count of the total number of mutant proteins is a better predictor of fitness than the number of proteins with variants scored as deleterious by PROVEAN. We also find that the sum of all mutant protein scores explains variation in fitness better than the number of mutant proteins in one of the data sets. We discuss the implications of these results for studies of populations in the wild.

Changes in ploidy are a significant type of genetic variation, describing the number of chromosome sets per cell. Ploidy evolves in natural populations, clinical populations, and lab experiments, particularly in unicellular fungi. Predicting how ploidy will evolve has proven difficult, despite a long history of theoretical work on this topic, as it is often unclear why one ploidy state outperforms another. Here, we review what is known about contemporary ploidy evolution in diverse fungal species through the lens of population genetics. As with typical genetic variants, ploidy evolution depends on the rate that new ploidy states arise by mutation, natural selection on alternative ploidy states, and random genetic drift. However, ploidy variation also has unique impacts on evolution, with the potential to alter chromosomal stability, the rate and patterns of point mutation, and the nature of selection on all loci in the genome. We discuss how ploidy evolution depends on these general and unique factors and highlight areas where additional experimental evidence is required to comprehensively explain the ploidy transitions observed in the field, the clinic, and the lab.

In sexual populations, the effectiveness of selection will depend on how gametes combine with respect to genetic quality. If gametes with deleterious alleles are likely to combine with one another, deleterious genetic variation can be more easily purged by selection. Assortative mating, where there is a positive correlation between parents in a phenotype of interest such as body size, is often observed in nature, but does not necessarily reveal how gametes ultimately combine with respect to genetic quality itself. We manipulated genetic quality in fruit fly populations using an inbreeding scheme designed to provide an unbiased measure of mating patterns. While inbred flies had substantially reduced reproductive success, their gametes did not combine with those of other inbred flies more often than expected by chance, indicating a lack of positive assortative mating. Instead, we detected a negative correlation in genetic quality between parents, i.e. disassortative mating, which diminished with age. This pattern is expected to reduce the genetic variance for fitness, diminishing the effectiveness of selection. We discuss how mechanisms of sexual selection could produce a pattern of disassortative mating. Our study highlights that sexual selection has the potential to either increase or decrease genetic load.

Despite decades of research, the factors that maintain genetic variation for fitness are poorly understood. It is unclear what fraction of the variance in a typical fitness component can be explained by mutation-selection balance (MSB) and whether fitness components differ in this respect. In theory, the level of standing variance in fitness due to MSB can be predicted using the rate of fitness decline under mutation accumulation, and this prediction can be directly compared to the standing variance observed. This approach allows for controlled statistical tests of the sufficiency of the MSB model, and could be used to identify traits or populations where genetic variance is maintained by other factors. For example, some traits may be influenced by sexually antagonistic balancing selection, resulting in an excess of standing variance beyond that generated by deleterious mutations. We describe the underlying theory and use it to test the MSB model for three traits in Drosophila melanogaster We find evidence for differences among traits, with MSB being sufficient to explain genetic variance in larval viability but not male mating success or female fecundity. Our results are consistent with balancing selection on sexual fitness components, and demonstrate the feasibility of rigorous statistical tests of the MSB model.