The contribution of spontaneous mutation to variation in environmental responses of Arabidopsis thaliana: responses to light

It has been hypothesized that new, spontaneous mutations tend to reduce fitness more severely in more stressful environments. To address this hypothesis, we grew plants representing 20 Arabidopsis thaliana mutation-accumulation (M-A) lines, advanced to generation 17, and their progenitor, in differing light conditions. The experiment was conducted in a greenhouse, and two treatments were used: full sun and shade, in which influx of red light was reduced relative to far-red. The shade treatment was considered the more stressful because mean absolute fitness was lower in that treatment, though not significantly so. Plants from generation 17 of M-A developed significantly faster than those from generation 0 in both treatments. A significant interaction between generation and treatment revealed that, counter to the hypothesis, M-A lines tended to have higher fitness on average relative to the progenitor in the shaded conditions, whereas, in full sun, the two generations were similar in fitness. A secondary objective of this experiment was to characterize the contribution of new mutations to genotype x environment interaction. We did not, however, detect a significant interaction between M-A line and treatment. Plots of the line-specific environmental responses indicate no tendency of new mutations to contribute to fitness trade-offs, between environments. They also do not support a model of conditionally deleterious mutation, in which a mutant reduces fitness only in a particular environment. These results suggest that interactions between genotype and light environment previously documented for A. thaliana are not explicable primarily as a consequence of steady input of spontaneous mutations having environment-specific effects.     

Fitness change in relation to mutation number in spontaneous mutation accumulation lines of Chlamydomonas reinhardtii

Although all genetic variation ultimately stems from mutations, their properties are difficult to study directly. Here, we used multiple mutation accumulation (MA) lines derived from five genetic backgrounds of the green algae Chlamydomonas reinhardtii that have been previously subjected to whole genome sequencing to investigate the relationship between the number of spontaneous mutations and change in fitness from a nonevolved ancestor. MA lines were on average less fit than their ancestors and we detected a significantly negative correlation between the change in fitness and the total number of accumulated mutations in the genome. Likewise, the number of mutations located within coding regions significantly and negatively impacted MA line fitness. We used the fitness data to parameterize a maximum likelihood model to estimate discrete categories of mutational effects, and found that models containing one to two mutational effect categories (one neutral and one deleterious category) fitted the data best. However, the best‐fitting mutational effects models were highly dependent on the genetic background of the ancestral strain.     

Fitness decline in spontaneous mutation accumulation lines of Caenorhabditis elegans with varying effective population sizes

The rate and fitness effects of new mutations have been investigated by mutation accumulation (MA) experiments in which organisms are maintained at a constant minimal population size to facilitate the accumulation of mutations with minimal efficacy of selection. We evolved 35 MA lines of Caenorhabditis elegans in parallel for 409 generations at three population sizes (N = 1, 10, and 100), representing the first spontaneous long-term MA experiment at varying population sizes with corresponding differences in the efficacy of selection. Productivity and survivorship in the N = 1 lines declined by 44% and 12%, respectively. The average effects of deleterious mutations in N = 1 lines are estimated to be 16.4% for productivity and 11.8% for survivorship. Larger populations (N = 10 and 100) did not suffer a significant decline in fitness traits despite a lengthy and sustained regime of consecutive bottlenecks exceeding 400 generations. Together, these results suggest that fitness decline in very small populations is dominated by mutations with large deleterious effects. It is possible that the MA lines at larger population sizes contain a load of cryptic deleterious mutations of small to moderate effects that would be revealed in more challenging environments.     

A comprehensive model of mutations affecting fitness and inferences for Arabidopsis thaliana

As the ultimate source of genetic variation, spontaneous mutation is essential to evolutionary change. Theoretical studies over several decades have revealed the dependence of evolutionary consequences of mutation on specific mutational properties, including genomic mutation rates, U, and the effects of newly arising mutations on individual fitness, s. The recent resurgence of empirical effort to infer these properties for diverse organisms has not achieved consensus. Estimates, which have been obtained by methods that assume mutations are unidirectional in their effects on fitness, are imprecise. Both because a general approach must allow for occurrence of fitness-enhancing mutations, even if these are rare, and because recent evidence demands it, we present a new method for inferring mutational parameters. For the distribution of mutational effects, we retain Keightley's assumption of the gamma distribution, to take advantage of the flexibility of its shape. Because the conventional gamma is one sided, restricting it to unidirectional effects, we include an additional parameter, rho, as an amount it is displaced from zero. Estimation is accomplished by Markov chain Monte Carlo maximum likelihood. Through a limited set of simulations, we verify the accuracy of this approach. We apply it to analyze data on two reproductive fitness components from a 17-generation mutation-accumulation study of a Columbia accession of Arabidopsis thaliana in which 40 lines sampled in three generations were assayed simultaneously. For these traits, U approximately/= 0.1-0.2, with distributions of mutational effects broadly spanning zero, such that roughly half the mutations reduce reproductive fitness. One evolutionary consequence of these results is lower extinction risks of small populations of A. thaliana than expected from the process of mutational meltdown. A comprehensive view of the evolutionary consequences of mutation will depend on quantitatively accounting for fitness-enhancing, as well as fitness-reducing, mutations.     

Fitness effects of mutation: testing genetic redundancy in Arabidopsis thaliana

Screens of organisms with disruptive mutations in a single gene often fail to detect phenotypic consequences for the majority of mutants. One explanation for this phenomenon is that the presence of paralogous loci provides genetic redundancy. However, it is also possible that the assayed traits are affected by few loci, that effects could be subtle or that phenotypic effects are restricted to certain environments. We assayed a set of T‐DNA insertion mutant lines of Arabidopsis thaliana to determine the frequency with which mutation affected fitness‐related phenotypes. We found that between 8% and 42% of the assayed lines had altered fitness from the wild type. Furthermore, many of these lines exhibited fitness greater than the wild type. In a second experiment, we grew a subset of the lines in multiple environments and found whether a T‐DNA insert increased or decreased fitness traits depended on the assay environment. Overall, our evidence contradicts the hypothesis that genetic redundancy is a common phenomenon in A. thaliana for fitness traits. Evidence for redundancy from prior screens of knockout mutants may often be an artefact of the design of the phenotypic assays which have focused on less complex phenotypes than fitness and have used single environments. Finally, our study adds to evidence that beneficial mutations may represent a significant component of the mutational spectrum of A. thaliana.     

Field measurements of genotype by environment interaction for fitness caused by spontaneous mutations in Arabidopsis thaliana

As the ultimate source of genetic diversity, spontaneous mutation is critical to the evolutionary process. The fitness effects of spontaneous mutations are almost always studied under controlled laboratory conditions rather than under the evolutionarily relevant conditions of the field. Of particular interest is the conditionality of new mutations—that is, is a new mutation harmful regardless of the environment in which it is found? In other words, what is the extent of genotype–environment interaction for spontaneous mutations? We studied the fitness effects of 25 generations of accumulated spontaneous mutations in Arabidopsis thaliana in two geographically widely separated field environments, in Michigan and Virginia. At both sites, mean total fitness of mutation accumulation lines exceeded that of the ancestors, contrary to the expected decrease in the mean due to new mutations but in accord with prior work on these MA lines. We observed genotype–environment interactions in the fitness effects of new mutations, such that the effects of mutations in Michigan were a poor predictor of their effects in Virginia and vice versa. In particular, mutational variance for fitness was much larger in Virginia compared to Michigan. This strong genotype–environment interaction would increase the amount of genetic variation maintained by mutation‐selection balance.     

Quantifying natural seasonal variation in mutation parameters with mutation accumulation lines

Mutations create novel genetic variants, but their contribution to variation in fitness and other phenotypes may depend on environmental conditions. Furthermore, natural environments may be highly heterogeneous. We assessed phenotypes associated with survival and reproductive success in over 30,000 plants representing 100 mutation accumulation lines of Arabidopsis thaliana across four temporal environments at a single field site. In each of the four assays, environmental variance was substantially larger than mutational variance. For some traits, whether mutational variance was significantly varied between seasons. The founder genotype had mean trait values near the mean of the distribution of the mutation accumulation lines in all field experiments. New mutations also contributed more phenotypic variation than would be predicted, given phenotypic and sequence‐level divergence among natural populations of A. thaliana. The combination of large environmental variance with a mean effect of mutation near zero suggests that mutations could contribute substantially to standing genetic variation.     

The contribution of spontaneous mutation to variation in environmental response in Arabidopsis thaliana: responses to nutrients

Abstract Although the evolutionary importance of spontaneous mutation is evident, its contribution to the evolution of ecological specificity remains unclear, because the environmental sensitivity of effects of new mutations has received little empirical attention. To address this issue, we report a greenhouse in which we grew plants from 20 mutation-accumulation (MA) lines, advanced by selfing and single-seed descent from a single common founder to generation 17, as well as plants from five lines representing the founder, in high and low nutrient conditions. We examined 11 traits throughout life history, including germination, survivorship, bolting date, flowering date, leaf number, leaf size, early and late height, mean fruit size, total seed weight, and reproductive biomass. Comparison of trait means between the two generations did not support the commonly held view that new mutations affecting fitness in these MA lines are strongly biased toward deleterious effects. We detected significant variance among MA lines for one fitness component, mean fruit size, but we did not detect a significant contribution of mutations accumulated in these MA lines to genotype by environment interaction (GEI). These results suggest that other evolutionary mechanisms play a more important role than spontaneous mutation alone in establishing the GEI found for wild collections and lab accessions of Arabidopsis thaliana in previous studies.     

Fitness decline under osmotic stress in Caenorhabditis elegans populations subjected to spontaneous mutation accumulation at varying population sizes

The consequences of mutations for population fitness depends on their individual selection coefficients and the effective population size. An earlier study of Caenorhabditis elegans spontaneous mutation accumulation lines evolved for 409 generations at three population sizes found that N_e  = 1 populations declined significantly in fitness whereas the fitness of larger populations (N_e  = 5, 50) was indistinguishable from the ancestral control under benign conditions. To test if larger MA populations harbor a load of cryptic deleterious mutations that are obscured under benign laboratory conditions, we measured fitness under osmotic stress via exposure to hypersaline conditions. The fitness of N_e  = 1 lines exhibited a further decline under osmotic stress compared to benign conditions. However, the fitness of larger populations remained indistinguishable from that of the ancestral control. The average effects of deleterious mutations in N_e  = 1 lines were estimated to be 22% for productivity and 14% for survivorship, exceeding values previously detected under benign conditions. Our results suggest that fitness decline is due to large effect mutations that are rapidly removed via selection even in small populations, with implications for conservation practices. Genetic stochasticity may not be as potent and immediate a threat to the persistence of small populations as other demographic and environmental stochastic factors.     

The fitness effects of spontaneous mutations in Caenorhabditis elegans

Abstract. Spontaneous mutation to mildly deleterious alleles has emerged as a potentially unifying component of a variety of observations in evolutionary genetics and molecular evolution. However, the biological significance of hypotheses based on mildly deleterious mutation depends critically on the rate at which new mutations arise and on their average effects. A long-term mutation-accumulation experiment with replicate lines of the nematode Caenorhabditis elegans maintained by single-progeny descent indicates that recurrent spontaneous mutation causes approximately 0.1% decline in fitness per generation, which is about an order of magnitude less than that suggested by previous studies with Drosophila . Two rather different approaches, Bateman-Mukai and maximum likelihood, suggest that this observation, along with the observed rate of increase in the variance of fitness among lines, is consistent with a genomic deleterious mutation rate for fitness of approximately 0.03 per generation and with an average homozygous effect of approximately 12%. The distribution of mutational effects for fitness appears to have a relatively low coefficient of variation, being no more extreme than expected for a negative exponential, and for one composite fitness measure (total progeny production) approaches constancy of effects. These results are derived from assays in a benign environment. At stressful temperatures, estimates of the genomic deleterious mutation rate (for genes expressed at such temperatures) is sixfold lower, whereas those for the average homozygous effect is approximately eightfold higher. Our results are reasonably compatible with existing estimates for flies, when one considers the differences between these species in the number of germ-line cell divisions per generation and the magnitude of transposable element activity.     

Flowering time QTL in natural populations of Arabidopsis thaliana and implications for their adaptive value

The genetic basis of phenotypic traits is of great interest to evolutionary biologists, but their contribution to adaptation in nature is often unknown. To determine the genetic architecture of flowering time in ecologically relevant conditions, we used a recombinant inbred line population created from two locally adapted populations of Arabidopsis thaliana from Sweden and Italy. Using these RILs, we identified flowering time QTL in growth chambers that mimicked the natural temperature and photoperiod variation across the growing season in each native environment. We also compared the genomic locations of flowering time QTL to those of fitness (total fruit number) QTL from a previous three‐year field study. Ten total flowering time QTL were found, and in all cases, the Italy genotype caused early flowering regardless of the conditions. Two QTL were consistent across chamber environments, and these had the largest effects on flowering time. Five of the fitness QTL colocalized with flowering time QTL found in the Italy conditions, and in each case, the local genotype was favoured. In contrast, just two flowering time QTL found in the Sweden conditions colocalized with fitness QTL and in only one case was the local genotype favoured. This implies that flowering time may be more important for adaptation in Italy than Sweden. Two candidate genes (FLC and VIN3) underlying the major flowering time QTL found in the current study are implicated in local adaptation.     

Analysis of the leaf metabolome in Arabidopsis thaliana mutation accumulation lines reveals association of metabolic disruption and fitness consequence

Evolutionary Ecology - Understanding the mechanisms by which mutations affect fitness and the distribution of mutational effects are central goals in evolutionary biology. Mutation accumulation...     

Evolution of interspecies unilateral incompatibility in the relatives of Arabidopsis thaliana

The evolutionary concurrence of intraspecies self‐incompatibility (SI) and explosive angiosperm radiation in the Cretaceous have led to the hypothesis that SI was one of the predominant drivers of rapid speciation in angiosperms. Interspecies unilateral incompatibility (UI) usually occurs when pollen from a self‐compatible (SC) species is rejected by the pistils of a SI species, while the reciprocal pollination is compatible (UC). Although this SI × SC type UI is most prevalent and viewed as a prezygotic isolation barrier to promote incipient speciation of angiosperms, comparative evidence to support such a role is lacking. We show that SI × SI type UI in SI species pairs is also common in the well‐characterized accessions representing the four major lineages of the Arabidopsis genus and is developmentally regulated. This allowed us to reveal a strong correlation between UI strength and species divergence in these representative accessions. In addition, analyses of a SC accession and the pseudo‐self‐compatible (psc) spontaneous mutant of Arabidopsis lyrata indicate that UI shares, at least, common pollen rejection pathway with SI. Furthermore, genetic and genomic analyses of SI × SI type UI in A. lyrata × A. arenosa species pair showed that two major‐effect quantitative trait loci are the stigma and pollen‐side determinant of UI, respectively, which could be involved in heterospecies pollen discrimination. By revealing a close link between UI and SI pathway, particularly between UI and species divergence in these representative accessions, our findings establish a connection between SI and speciation. Thus, the pre‐existence of SI system would have facilitated the evolution of UI and accordingly promote speciation.     

The effects of pathogen infection and mutation on life-history characters in Arabidopsis thaliana

The nature of the interaction among deleterious mutations is important to models in many areas of evolutionary biology. In addition, interactions between genetic and environmental factors may affect the predictions of such models. Individuals of unknown genotypes of Arabidopsis thaliana, ecotype Marburg, were exposed to five levels of chemical (EMS) mutagenesis and three levels of Pseudomonas syringae infection. Survival, growth and flowering characteristics of each individual were measured. The logarithm of fitness is expected to be a linear function of mutation number if mutations act independently. Furthermore, the expected number of mutations should be approximately a linear function of time of exposure to mutagen. Therefore, nonlinear effects of mutagen exposure on the logarithm of fitness characters would suggest epistasis between mutations. Similarly, if pathogen infection and mutation act independently of each other, their effects should be additive on a log scale. Statistical interactions between these factors would suggest they do not act independently; particularly, if highly mutated individuals suffer more when infected than do less mutated individuals, this suggests that pathogens and mutations act synergistically. Pseudomonas-infected individuals were shown to have an increased probability of flowering under conditions of short day length, but to ultimately produce fewer flowers than uninfected individuals. This suggests a plastic response to stress and, despite that response, an ultimately deleterious effect of infection on fitness. Leaf rosette growth was negatively and linearly related to the expected number of mutations, and the effects of mutation on different life-cycle stages appeared to be uncorrelated. No significant interactions between pathogen and mutation main effects were found. These results suggest that mutations act multiplicatively with each other and with pathogen infection in determining individual fitness.     

Inside the heads of David and Goliath: environmental effects on brain morphology among wild and growth-enhanced coho salmon Oncorhynchus kisutch

Transgenic and wild-type individual coho salmon Oncorhynchus kisutch were reared in hatchery and near-natural stream conditions and their brain and structure sizes were determined. Animals reared in the hatchery grew larger and developed larger brains, both absolutely and when controlling for body size. In both environments, transgenics developed relatively smaller brains than wild types. Further, the volume of the optic tectum of both genotypes was larger in the hatchery animals and the cerebellum of transgenics was smaller when reared in near-natural streams. Finally, wild types developed a markedly smaller telencephalon under hatchery conditions. It is concluded that, apart from the environment, genetic factors that modulate somatic growth rate also have a strong influence on brain size and structure.     

Characterization of Arabidopsis thaliana mismatch specific endonucleases: application to mutation discovery by TILLING in pea

Scanning DNA sequences for mutations and polymorphisms has become one of the most challenging, often expensive and time-consuming obstacles in many molecular genetic applications, including reverse genetic and clinical diagnostic applications. Enzymatic mutation detection methods are based on the cleavage of heteroduplex DNA at the mismatch sites. These methods are often limited by the availability of a mismatch-specific endonuclease, their sensitivity in detecting one allele in a pool of DNA and their costs. Here, we present detailed biochemical analysis of five Arabidopsis putative mismatch-specific endonucleases. One of them, ENDO1, is presented as the first endonuclease that recognizes and cleaves all types of mismatches with high efficiency. We report on a very simple protocol for the expression and purification of ENDO1. The ENDO1 system could be exploited in a wide range of mutation diagnostic tools. In particular, we report the use of ENDO1 for discovery of point mutations in the gibberellin 3beta-hydrolase gene of Pisum sativum. Twenty-one independent mutants were isolated, five of these were characterized and two new mutations affecting internodes length were identified. To further evaluate the quality of the mutant population we screened for mutations in four other genes and identified 5-21 new alleles per target. Based on the frequency of the obtained alleles we concluded that the pea population described here would be suitable for use in a large reverse-genetics project.     

Towards the genetic architecture of seed lipid biosynthesis and accumulation in Arabidopsis thaliana

We report the quantitative genetic analysis of seed oil quality and quantity in six Arabidopsis thaliana recombinant inbred populations, in which the parent accessions were from diverse geographical origins, and were selected on the basis of variation for seed oil content and lipid composition. Although most of the biochemical steps involved in lipid biosynthesis are known and the key genes have been identified, the regulation of the processes that results in the final oil composition and total amount is not understood. By using physically anchored markers it was possible to compare results across populations. A total of 219 quantitative trait loci (QTLs) were identified, of which 81 were significant at P<0.001. Some of these colocalise with QTLs identified previously, but many novel QTLs were also identified. The results highlight the importance of studying traits in multiple populations, which will lead to a better understanding of the contribution that natural variation makes to the genetic architecture of a phenotype.     

Effects of spontaneous mutation accumulation on sex ratio traits in a parasitoid wasp

Sex allocation theory has proved extremely successful at predicting when individuals should adjust the sex of their offspring in response to environmental conditions. However, we know rather little about the underlying genetics of sex ratio or how genetic architecture might constrain adaptive sex-ratio behavior. We examined how mutation influenced genetic variation in the sex ratios produced by the parasitoid wasp Nasonia vitripennis. In a mutation accumulation experiment, we determined the mutability of sex ratio, and compared this with the amount of genetic variation observed in natural populations. We found that the mutability (h(2)(m)) ranges from 0.001 to 0.002, similar to estimates for life-history traits in other organisms. These estimates suggest one mutation every 5-60 generations, which shift the sex ratio by approximately 0.01 (proportion males). In this and other studies, the genetic variation in N. vitripennis sex ratio ranged from 0.02 to 0.17 (broad-sense heritability, H(2)). If sex ratio is maintained by mutation-selection balance, a higher genetic variance would be expected given our mutational parameters. Instead, the observed genetic variance perhaps suggests additional selection against sex-ratio mutations with deleterious effects on other fitness traits as well as sex ratio (i.e., pleiotropy), as has been argued to be the case more generally.     

Genetic analysis of the physiological responses to low boron stress in Arabidopsis thaliana

Boron (B) is an essential micronutrient for higher plants. There is wide genetic variation in the response to B deficiency among plant species and cultivars. The objective of this study was to identify quantitative trait loci (QTL) that control B efficiency in natural Arabidopsis accessions. The B efficiency coefficient (BEC) and seed yield under low B conditions (SYLB) were investigated by solution culture in two separate experiments in an Arabidopsis recombinant inbred line (RIL) population. Both of the traits studied exhibited high transgressive variation in the RIL population, and, in total, five and three QTL were identified for BEC and SYLB, respectively. Three of the five QTL, including the QTL, AtBE1-2, that has a large effect on the BEC, were found at the interval of the corresponding QTL for SYLB in both experiments. The close genetic relationship between BEC and SYLB was further confirmed by conditional QTL mapping in the RIL population and unconditional QTL mapping in an AtBE1-2-segregated F(2) population. Epistatic interactions for the tested traits were analysed, and were found to be widespread in the detected QTL of Arabidopsis in the RIL population. Comparison of the QTL interval for B efficiency with reported B-related genes showed that 10 B-related genes, together with one BOR1 homolog (BOR5, At1g74810) were located in the QTL region of AtBE1-2. These results suggest that natural variation in B efficiency in Arabidopsis has a complex molecular basis. They also provide a basis for fine mapping and cloning of the B-efficiency genes, with the ultimate aim of discovering the physiological mechanism of action of the genes.