Heterogeneous evolution of microsatellites revealed by reconstruction of recent mutation history in an invasive apomictic snail, Potamopyrgus antipodarum

Heterogeneous patterns of microsatellite evolution present a major challenge for the development of mutation models, and an improved understanding of the determinants of variation in mutation rates and patterns among loci, alleles and taxa is required. A 19th Century bottleneck associated with the introduction of clones of the snail Potamopyrgus antipodarum to Britain presented an opportunity to reconstruct recent microsatellite evolution within the most common apomictic lineage. There was significant variation in both the number and step size of mutations among the seven loci studied. Patterns of mutability were consistent with higher mutation rates for di- than trinucleotides and for longer alleles at a locus. Mutation size was influenced in a more complex way, decreasing with relative allele length much more strongly for tri-, than dinucleotides. We found support for this latter, highly novel result in the literature via reanalysis of data in a recent genome-scan study of human microsatellites, which showed a similarly disparate pattern of length-dependence between di- and trinucleotides. In spite of the apomictic form of reproduction and an unusually strong excess of microsatellite contractions in P. antipodarum, there were notable similarities with mutation processes of human microsatellites, supporting the wider taxonomic generality of such evolutionary mechanisms.     

Mutating away from your enemies: The evolution of mutation rate in a host–parasite system

The rate at which mutations occur in nature is itself under natural selection. While a general reduction of mutation rates is advantageous for species inhabiting constant environments, higher mutation rates can be advantageous for those inhabiting fluctuating environments that impose on-going directional selection. Analogously, species involved in antagonistic co-evolutionary arms races, such as hosts and parasites, can also benefit from higher mutation rates. We use modifier theory, combined with simulations, to investigate the evolution of mutation rate in such a host–parasite system. We derive an expression for the evolutionary stable mutation rate between two alleles, each of whose fitness depends on the current genetic composition of the other species. Recombination has been shown to weaken the strength of selection acting on mutation modifiers, and accordingly, we find that the evolutionarily attracting mutation rate is lower when recombination between the selected and the modifier locus is high. Cyclical dynamics are potentially commonplace for loci governing antagonistic species interactions. We characterize the parameter space where such cyclical dynamics occur and show that the evolution of large mutation rates tends to inhibit cycling and thus eliminates further selection on modifiers of the mutation rate. We then find using computer simulations that stochastic fluctuations in finite populations can increase the size of the region where cycles occur, creating selection for higher mutation rates. We finally use simulations to investigate the model behaviour when there are more than two alleles, finding that the region where cycling occurs becomes smaller and the evolutionarily attracting mutation rate lower when there are more alleles.     

Premeiotic clusters of mutation and the cost of natural selection

Haldane stated that there is a cost of natural selection for new beneficial alleles to be substituted over time. Most of this cost, which leads to "genetic deaths," is in the early generations of the substitution process when the new allele is low in frequency. It depends on the initial frequency and dominance value, but not the selection coefficient, of the advantageous allele. There have been numerous suggestions on how to reduce the cost for preexisting genetic variation that goes from disadvantageous, or neutral, to advantageous with a change in the environment. However, the cost of natural selection for new alleles that arise by mutation is assumed to be high, based on the assumption that new mutant alleles arise in natural populations as single events [1/(2N) of the total alleles]. However, not all mutant alleles arise as single events. Premeiotic mutations occur frequently in individuals (germinal mosaics), giving rise to multiple copies of identical mutant alleles called a "cluster" (C) with an initial allele frequency of C/(2N) instead of 1/(2N). These clusters of new mutant alleles reduce the cost of natural selection in direct proportion to the relative size of the cluster. Hence new advantageous alleles that arise by mutation have the greatest chance of going to fixation if they occur in large clusters in small populations.     

Estimating the degree of saturation in mutant screens

Large-scale screens for loss-of-function mutants have played a significant role in recent advances in developmental biology and other fields. In such mutant screens, it is desirable to estimate the degree of "saturation" of the screen (i.e., what fraction of the possible target genes has been identified). We applied Bayesian and maximum-likelihood methods for estimating the number of loci remaining undetected in large-scale screens and produced credibility intervals to assess the uncertainty of these estimates. Since different loci may mutate to alleles with detectable phenotypes at different rates, we also incorporated variation in the degree of mutability among genes, using either gamma-distributed mutation rates or multiple discrete mutation rate classes. We examined eight published data sets from large-scale mutant screens and found that credibility intervals are much broader than implied by previous assumptions about the degree of saturation of screens. The likelihood methods presented here are a significantly better fit to data from published experiments than estimates based on the Poisson distribution, which implicitly assumes a single mutation rate for all loci. The results are reasonably robust to different models of variation in the mutability of genes. We tested our methods against mutant allele data from a region of the Drosophila melanogaster genome for which there is an independent genomics-based estimate of the number of undetected loci and found that the number of such loci falls within the predicted credibility interval for our models. The methods we have developed may also be useful for estimating the degree of saturation in other types of genetic screens in addition to classical screens for simple loss-of-function mutants, including genetic modifier screens and screens for protein-protein interactions using the yeast two-hybrid method.     

Detecting the molecular signature of social conflict: theory and a test with bacterial quorum sensing genes

Extending social evolution theory to the molecular level opens the door to an unparalleled abundance of data and statistical tools for testing alternative hypotheses about the long-term evolutionary dynamics of cooperation and conflict. To this end, we take a collection of known sociality genes (bacterial quorum sensing [QS] genes), model their evolution in terms of patterns that are detectable using gene sequence data, and then test model predictions using available genetic data sets. Specifically, we test two alternative hypotheses of social conflict: (1) the "adaptive" hypothesis that cheaters are maintained in natural populations by frequency-dependent balancing selection as an evolutionarily stable strategy and (2) the "evolutionary null" hypothesis that cheaters are opposed by purifying kin selection yet exist transiently because of their recurrent introduction into populations by mutation (i.e., kin selection-mutation balance). We find that QS genes have elevated within- and among-species sequence variation, nonsignificant signatures of natural selection, and putatively small effect sizes of mutant alleles, all patterns predicted by our evolutionary null model but not by the stable cheater hypothesis. These empirical findings support our theoretical prediction that QS genes experience relaxed selection due to nonclonality of social groups, conditional expression, and the individual-level advantage enjoyed by cheaters. Furthermore, cheaters are evolutionarily transient, persisting in populations because of their recurrent introduction by mutation and not because they enjoy a frequency-dependent fitness advantage.     

Allele multiplicity in simple Mendelian disorders.

A model of selection involving two selectively equivalent classes of alleles at a locus is considered. One class consists of normal alleles A1, A2, A3,. . .; the other class consists of detrimental alleles a1, a2, a3, . . . . Mutation within and between allelic classes can occur without restriction, but selection operates in such a way as to maintain an approximately constant overall frequency of A-type and a-type alleles is derived, and it is shown that the distribution of allele frequencies in a sample of detrimental alleles depends on the forward (A to a) mutation rate but not on the selection coefficient, degree of dominance, or mutation rate among a-type alleles. Recurrent mutation therefore generates allelic multiplicity among detrimental alleles, and this is discussed in the context of clinical heterogeneity in simple Mendelian disorders.     

Genetic instability at the opaque-2 locus of maize

Summary A case of genetic variegation discovered at the opaque-2 locus of maize that includes a two-element system with a receptor and regulatory element is described. The somatic mutability depends on the existence of two genetic factors: a responsive allele (with receptor element), o2m(r), and a regulatory element, Bg, that induces mutability of o2m(r). In the absence of Bg, o2m(r) is indistiguishable from the recessive alleles of the O2 locus; in the presence of the regulatory element, o2m(r) mutates giving rise to sectors of flint-like endosperm in an opaque back-ground. The regulatory element Bg may be located independently or at the controlled locus. The genetic properties of the new system, somatic mutability, transposition, existence of different patterns of mutability, are apparently similar to those previously described in maize for the classical systems of controlling elements. In addition, the recovery of the o2 mutability from crosses between spontaneous o2 alleles suggests that transposable genetic elements may be involved in the origin of natural mutability.     

Bias in the introduction of variation as an orienting factor in evolution

SUMMARY According to New Synthesis doctrine, the direction of evolution is determined by selection and not by "internal causes" that act by way of propensities of variation. This doctrine rests on the theoretical claim that because mutation rates are small in comparison to selection coefficients, mutation is powerless to overcome opposing selection. Using a simple population-genetic model, this claim is shown to depend on assuming the prior availability of variation, so that mutation may act only as a "pressure" on the frequencies of existing alleles, and not as the evolutionary process that introduces novelty. As shown here, mutational bias in the introduction of novelty can strongly influence the course of evolution, even when mutation rates are small in comparison to selection coefficients. Recognizing this mode of causation provides a distinct mechanistic basis for an "internalist" approach to determining the contribution of mutational and developmental factors to evolutionary phenomena such as homoplasy, parallelism, and directionality.     

Is a mutator analogous to the Ig hypermutator of the sheep ileal Peyer's patch active on MHC class I genes in the germ line?

 Polymorphic sequence variation in the peptide-binding domains of MHC class I molecules appears to have been driven largely by the constructive action of natural selection on the specificity of the peptide-binding groove. Similar features are displayed by the variable domains of immunoglobulins generated in the sheep ileal Peyer's patch, but in this case there is evidence that the action of a targeted hypermutator acting on a selected substrate rather than antigen-driven selection is responsible for the pattern of variation in the system. Such a hypermutator acting in the germ line would not only mimic the action of natural selection but also, by convergent mutation, generate similar patterns of variation in unrelated alleles that could be interpreted as evidence for short-tract gene conversion. We analyzed human class I MHC alleles in the light of these data, but failed to find evidence of the action of a similar hypermutator. A search for other mutationally driven patterns of variation also failed, even in hypervariable residues from parsimonious phylogenies. Single-nucleotide variation at these residues is also frequent in recent allelic variants, but the data are as consistent with short-tract gene conversion as with base mutation. We conclude that the patterns of allelic variation in MHC molecules are not driven by mutational pressure, but rather by conventional mutational processes, accompanied by short-tract gene conversion and intense natural selection. Received: 6 October 1999 / Revised: 30 December 1999     

Risk estimation based on germ-cell mutations in animals

The set of mouse germ cell mutation rate results following spermatogonial exposure to high dose rate irradiation have been presented as the most relevant experimental results upon which to extrapolate the expected genetic risk of offspring of the survivors of the Hiroshima and Nagasaki atomic bombings. Results include mutation rates to recessive specific-locus, dominant cataract, protein-charge, and enzyme-activity alleles. The mutability as determined by the various genetic end points differed: the mutation rates to recessive specific-locus alleles and enzyme-activity alleles were similar and greater than the mutation rates to dominant cataract and protein-charge alleles. It is argued that the type of mutation event scored by a particular test will determine the mutability of the genetic end point screened. When the loss of functional gene product can be scored in a particular mutation test, as in the recessive specific-locus and enzyme-activity tests, a wide spectrum of DNA alterations may result in a loss of and a higher mutation rate is observed. When an altered gene product is scored, as in the dominant cataract and protein-charge tests, a narrower spectrum of DNA alterations is screened and a lower mutation rate is observed. The radiation doubling dose, defined as the dose that induces as many mutations as occur spontaneously per generation, was shown to be four times higher in the dominant cataract test than the specific-locus test. These results indicate that to extrapolate to genetic risks in humans using the doubling-dose method, the extrapolation must be based on experimental mutation rate results for the same genetic end point.(ABSTRACT TRUNCATED AT 250 WORDS)     

Some features of the mutability of bacteria during nonlethal selection

We describe the mutability of the Trp(-) chromosomal +1 frameshift mutation trpE7999 during nonlethal selection, finding that the appearance of Trp(+) revertants behaves similarly to that of episomal Lac(+) revertants. In addition, we show that a feature of the Lac(+) and Trp(+) mutability is the accumulation of Trp(+) and Lac(+) revertants with additional unselected mutations, most of which are not due to heritable mutators. The cells undergoing nonlethal selection apparently experience an epigenetic change resulting in a subset of bacteria with elevated mutability that often remain hypermutable for the duration of selection. The epigenetic change provoked by nonlethal selection appears to be mediated by a unique function provided by the F'128 episome.     

A general model to explore complex dominance patterns in plant sporophytic self-incompatibility systems

We developed a general model of sporophytic self-incompatibility under negative frequency-dependent selection allowing complex patterns of dominance among alleles. We used this model deterministically to investigate the effects on equilibrium allelic frequencies of the number of dominance classes, the number of alleles per dominance class, the asymmetry in dominance expression between pollen and pistil, and whether selection acts on male fitness only or both on male and on female fitnesses. We show that the so-called "recessive effect" occurs under a wide variety of situations. We found emerging properties of finite population models with several alleles per dominance class such as that higher numbers of alleles are maintained in more dominant classes and that the number of dominance classes can evolve. We also investigated the occurrence of homozygous genotypes and found that substantial proportions of those can occur for the most recessive alleles. We used the model for two species with complex dominance patterns to test whether allelic frequencies in natural populations are in agreement with the distribution predicted by our model. We suggest that the model can be used to test explicitly for additional, allele-specific, selective forces.     

A general population genetic framework for antagonistic selection that accounts for demography and recurrent mutation

Antagonistic selection--where alleles at a locus have opposing effects on male and female fitness ("sexual antagonism") or between components of fitness ("antagonistic pleiotropy")--might play an important role in maintaining population genetic variation and in driving phylogenetic and genomic patterns of sexual dimorphism and life-history evolution. While prior theory has thoroughly characterized the conditions necessary for antagonistic balancing selection to operate, we currently know little about the evolutionary interactions between antagonistic selection, recurrent mutation, and genetic drift, which should collectively shape empirical patterns of genetic variation. To fill this void, we developed and analyzed a series of population genetic models that simultaneously incorporate these processes. Our models identify two general properties of antagonistically selected loci. First, antagonistic selection inflates heterozygosity and fitness variance across a broad parameter range--a result that applies to alleles maintained by balancing selection and by recurrent mutation. Second, effective population size and genetic drift profoundly affect the statistical frequency distributions of antagonistically selected alleles. The "efficacy" of antagonistic selection (i.e., its tendency to dominate over genetic drift) is extremely weak relative to classical models, such as directional selection and overdominance. Alleles meeting traditional criteria for strong selection (N(e)s > 1, where N(e) is the effective population size, and s is a selection coefficient for a given sex or fitness component) may nevertheless evolve as if neutral. The effects of mutation and demography may generate population differences in overall levels of antagonistic fitness variation, as well as molecular population genetic signatures of balancing selection.     

Natural Selection and the Frequency Distributions of ``silent' DNA Polymorphism in Drosophila

In Escherichia coli, Saccharomyces cerevisiae, and Drosophila melanogaster, codon bias may be maintained by a balance among mutation pressure, genetic drift, and natural selection favoring translationally superior codons. Under such an evolutionary model, silent mutations fall into two fitness categories: preferred mutations that increase codon bias and unpreferred changes in the opposite direction. This prediction can be tested by comparing the frequency spectra of synonymous changes segregating within populations; natural selection will elevate the frequencies of advantageous mutations relative to that of deleterious changes. The frequency distributions of preferred and unpreferred mutations differ in the predicted direction among 99 alleles of two D. pseudoobscura genes and five alleles of eight D. simulans genes. This result confirms the existence of fitness classes of silent mutations. Maximum likelihood estimates suggest that selection intensity at silent sites is, on average, very weak in both D. pseudoobscura and D. simulans (|N(e)s| & 1). Inference of evolutionary processes from within-species sequence variation is often hindered by the assumption of a stationary frequency distribution. This assumption can be avoided when identifying the action of selection and tested when estimating selection intensity.     

Experimental evolution and the dynamics of genomic mutation rate modifiers

Because genes that affect mutation rates are themselves subject to mutation, mutation rates can be influenced by natural selection and other evolutionary forces. The population genetics of mutation rate modifier alleles has been a subject of theoretical interest for many decades. Here, we review experimental contributions to our understanding of mutation rate modifier dynamics. Numerous evolution experiments have shown that mutator alleles (modifiers that elevate the genomic mutation rate) can readily rise to high frequencies via genetic hitchhiking in non-recombining microbial populations. Whereas these results certainly provide an explanatory framework for observations of sporadically high mutation rates in pathogenic microbes and in cancer lineages, it is nonetheless true that most natural populations have very low mutation rates. This raises the interesting question of how mutator hitchhiking is suppressed or its phenotypic effect reversed in natural populations. Very little experimental work has addressed this question; with this in mind, we identify some promising areas for future experimental investigation.     

The microbiology of mutability

Bacteria possessing elevated spontaneous mutation rates are prevalent in certain environments, which is a paradox because most mutations are deleterious. For example, cells with defects in the methyl-directed mismatch repair (MMR) system, termed mutators or hypermutators, are overrepresented in populations of bacterial pathogens, with the mutator trait hypothesized to be advantageous in the changing host enviroments faced during colonization and establishment of chronic infections. Error-prone DNA polymerases, such as polIV and polV, function in translesion DNA synthesis, a DNA damage response that ensures genome integrity with a cost of increased mutation. While the biochemical aspects of these mutability pathways are well understood, the biological impacts have received less attention. Here, an examination of bacterial mutability systems and specifically the ecological and evolutionary context resulting in the selection of these systems is carried out.     

Evaluation of the flanking nucleotide sequences of sarcomeric hypertrophic cardiomyopathy substitution mutations

Hypertrophic cardiomyopathy (HCM) is a familial myocardial disease with a prevalence of 1 in 500. More than 400 causative mutations have been identified in 13 sarcomeric and myofilament related genes, 350 of these are substitution mutations within eight sarcomeric genes. Within a population, examples of recurring identical disease causing mutations that appear to have arisen independently have been noted as well as those that appear to have been inherited from a common ancestor. The large number of novel HCM mutations could suggest a mechanism of increased mutability within the sarcomeric genes. The objective of this study was to evaluate the most commonly reported HCM genes, beta myosin heavy chain (MYH7), myosin binding protein C, troponin I, troponin T, cardiac regulatory myosin light chain, cardiac essential myosin light chain, alpha tropomyosin and cardiac alpha-actin for sequence patterns surrounding the substitution mutations that may suggest a mechanism of increased mutability. The mutations as well as the 10 flanking nucleotides were evaluated for frequency of di-, tri- and tetranucleotides containing the mutation as well as for the presence of certain tri- and tetranculeotide motifs. The most common substitutions were guanine (G) to adenine (A) and cytosine (C) to thymidine (T). The CG dinucleotide had a significantly higher relative mutability than any other dinucleotide (p<0.05). The relative mutability of each possible trinucleotide and tetranucleotide sequence containing the mutation was calculated; none were at a statistically higher frequency than the others. The large number of G to A and C to T mutations as well as the relative mutability of CG may suggest that deamination of methylated CpG is an important mechanism for mutation development in at least some of these cardiac genes.     

Single nucleotide polymorphism-based validation of exonic splicing enhancers

Because deleterious alleles arising from mutation are filtered by natural selection, mutations that create such alleles will be underrepresented in the set of common genetic variation existing in a population at any given time. Here, we describe an approach based on this idea called VERIFY (variant elimination reinforces functionality), which can be used to assess the extent of natural selection acting on an oligonucleotide motif or set of motifs predicted to have biological activity. As an application of this approach, we analyzed a set of 238 hexanucleotides previously predicted to have exonic splicing enhancer (ESE) activity in human exons using the relative enhancer and silencer classification by unanimous enrichment (RESCUE)-ESE method. Aligning the single nucleotide polymorphisms (SNPs) from the public human SNP database to the chimpanzee genome allowed inference of the direction of the mutations that created present-day SNPs. Analyzing the set of SNPs that overlap RESCUE-ESE hexamers, we conclude that nearly one-fifth of the mutations that disrupt predicted ESEs have been eliminated by natural selection (odds ratio = 0.82 +/- 0.05). This selection is strongest for the predicted ESEs that are located near splice sites. Our results demonstrate a novel approach for quantifying the extent of natural selection acting on candidate functional motifs and also suggest certain features of mutations/SNPs, such as proximity to the splice site and disruption or alteration of predicted ESEs, that should be useful in identifying variants that might cause a biological phenotype.     

Impact of Genetic Background on Allele Selection in a Highly Mutable Candida albicans Gene,PNG2

In many microbes rapid mutation of highly mutable contingency genes continually replenishes a pool of variant alleles from which the most suitable are selected, assisting in rapid adaptation and evasion of the immune response. In some contingency genes mutability is achieved through DNA repeats within the coding region. The fungal human pathogen Candida albicans has 2600 repeat-containing ORFs. For those investigated (ALS genes, HYR1, HYR2, CEK1, RLM1) many protein variants with differing amino acid repeat regions exist, as expected for contingency genes. However, specific alleles dominate in different clades, which is unexpected if allele variation is used for short-term adaptation. Generation of new alleles of repeat-containing C. albicans ORFs has never been observed directly. Here we present evidence for restrictions on the emergence of new alleles in a highly mutable C. albicans repeat-containing ORF, PNG2, encoding a putative secreted or cell surface glycoamidase. In laboratory cultures new PNG2 alleles arose at a rate of 2.8×10⁻⁵ (confidence interval 3.3×10⁻⁶−9. 9×10⁻⁵) per cell per division, comparable to rates measured for contingency genes. Among 80 clinical isolates 17 alleles of different length and 23 allele combinations were distinguishable; sequence differences between repeat regions of identical size suggest the existence of 36 protein variants. Specific allele combinations predominated in different genetic backgrounds, as defined by DNA fingerprinting and multilocus sequence typing. Given the PNG2 mutation rate, this is unexpected, unless in different genetic backgrounds selection favors different alleles. Specific alleles or allele combinations were not preferentially associated with C. albicans isolates from particular body sites or geographical regions. Our results suggest that the mutability of PNG2 is not used for short-term adaptation or evasion of the immune system. Nevertheless the large number of alleles observed indicates that mutability of PNG2 may assist C. albicans strains from different genetic backgrounds optimize their interaction with the host in the long term.     

Molecular evolution of two linked genes, Est-6 and Sod, in Drosophila melanogaster.

We have obtained 15 sequences of Est-6 from a natural population of Drosophila melanogaster to test whether linkage disequilibrium exists between Est-6 and the closely linked Sod, and whether natural selection may be involved. An early experiment with allozymes had shown linkage disequilibrium between these two loci, while none was detected between other gene pairs. The Sod sequences for the same 15 haplotypes were obtained previously. The two genes exhibit similar levels of nucleotide polymorphism, but the patterns are different. In Est-6, there are nine amino acid replacement polymorphisms, one of which accounts for the S-F allozyme polymorphism. In Sod, there is only one replacement polymorphism, which corresponds to the S-F allozyme polymorphism. The transversion/transition ratio is more than five times larger in Sod than in Est-6. At the nucleotide level, the S and F alleles of Est-6 make up two allele families that are quite different from each other, while there is relatively little variation within each of them. There are also two families of alleles in Sod, one consisting of a subset of F alleles, and the other consisting of another subset of F alleles, designed F(A), plus all the S alleles. The Sod F(A) and S alleles are completely or nearly identical in nucleotide sequence, except for the replacement mutation that accounts for the allozyme difference. The two allele families have independent evolutionary histories in the two genes. There are traces of statistically significant linkage disequilibrium between the two genes that, we suggest, may have arisen as a consequence of selection favoring one particular sequence at each locus.