"Reverse ecology" and the power of population genomics

Rapid and inexpensive sequencing technologies are making it possible to collect whole genome sequence data on multiple individuals from a population. This type of data can be used to quickly identify genes that control important ecological and evolutionary phenotypes by finding the targets of adaptive natural selection, and we therefore refer to such approaches as "reverse ecology." To quantify the power gained in detecting positive selection using population genomic data, we compare three statistical methods for identifying targets of selection: the McDonald-Kreitman test, the mkprf method, and a likelihood implementation for detecting d(N)/d(S) > 1. Because the first two methods use polymorphism data we expect them to have more power to detect selection. However, when applied to population genomic datasets from human, fly, and yeast, the tests using polymorphism data were actually weaker in two of the three datasets. We explore reasons why the simpler comparative method has identified more genes under selection, and suggest that the different methods may really be detecting different signals from the same sequence data. Finally, we find several statistical anomalies associated with the mkprf method, including an almost linear dependence between the number of positively selected genes identified and the prior distributions used. We conclude that interpreting the results produced by this method should be done with some caution.     

The relationship between microsatellite polymorphism and recombination hot spots in the human genome

Although previous studies have failed to detect an association between microsatellite polymorphism and broadscale recombination rates in the human genome, there are several possible reasons why such a relationship could exist. For instance, there might be a direct link if recombination is mutagenic to microsatellite sequences or if polymorphic microsatellites act as recombination signals. Alternatively, recombination could exert an indirect effect by uncoupling of natural selection at linked loci, promoting polymorphism. As recombination is concentrated in narrow hotspot regions in the human genome, we investigated the relationship between microsatellite polymorphism and recombination hot spots. By using data from a common allele frequency database, we found several polymorphism estimates to be similar for hot spots and the genomic average. However, this is likely explained by an ascertainment bias because markers with high polymorphism information content are usually selected for genotyping in human populations and pedigrees. In contrast, by using an unbiased set of shotgun sequence data, we found an excess of microsatellite polymorphism in recombination hot spots of 14%. However, when other genomic variables are taken into account in a generalized model and using wavelet analysis, the effect is no longer detectable and the only firm predictor of microsatellite polymorphism is the incidence of SNPs and indels. One possible neutral explanation to these observations is that there is a common denominator affecting the local rate of mutation in unique as well as in repetitive DNA, for example, base composition.     

Relative roles of mutation and recombination in generating allelic polymorphism at an MHC class II locus in Peromyscus maniculatus

The MHC class II loci encoding cell surface antigens exhibit extremely high allelic polymorphism. There is considerable uncertainty in the literature over the relative roles of recombination and de novo mutation in generating this diversity. We studied class II sequence diversity and allelic polymorphism in two populations of Peromyscus maniculatus, which are among the most widespread and abundant mammals of North America. We find that intragenic recombination (or gene conversion) has been the predominant mode for the generation of allelic polymorphism in this species, with the amount of population recombination per base pair exceeding mutation by at least an order of magnitude during the history of the sample. Despite this, patchwork motifs of sites with high linkage disequilibrium are observed. This does not appear to be consistent with the much larger amount of recombination versus mutation in the history of the sample, unless the recombination rate is highly non-uniform over the sequence or selection maintains certain sites in linkage disequilibrium. We conclude that selection is most likely to be responsible for preserving sequence motifs in the presence of abundant recombination.     

Persistent genomic instability in the yeast Saccharomyces cerevisiae induced by ionizing radiation and DNA-damaging agents

A "hypermutable" genome is a common characteristic of cancer cells, and it may contribute to the progressive accumulation of mutations required for the development of cancer. It has been reported that mammalian cells surviving exposure to gamma radiation display several highly persistent genomic instability phenotypes which may reflect a hypermutability similar to that seen in cancer. These phenotypes include an increased mutation frequency and a decreased plating efficiency, and they continue to be observed many generations after the radiation exposure. The underlying causes of this genomic instability have not been fully determined. We show here that exposure to gamma radiation and other DNA-damaging treatments induces a similar genomic instability in the yeast Saccharomyces cerevisiae. A dose-dependent increase in intrachromosomal recombination was observed in cultures derived from cells surviving gamma irradiation as many as 50 generations after the exposure. Increased forward mutation frequencies and low colony-forming efficiencies were also observed. Persistently elevated recombination frequencies in haploid cells were dominant after these cells were mated to nonirradiated partners, and the elevated recombination phenotype was also observed after treatment with the DNA-damaging agents ultraviolet light, hydrogen peroxide, and ethyl methanesulfonate. Radiation-induced genomic instability in yeast may represent a convenient model for the hypermutability observed in cancer cells.     

Linkage disequilibria and the site frequency spectra in the su(s) and su(w(a)) regions of the Drosophila melanogaster X chromosome

Over the last decade, surveys of DNA sequence variation in natural populations of several Drosophila species and other taxa have established that polymorphism is reduced in genomic regions characterized by low rates of crossing over per physical length. Parallel studies have also established that divergence between species is not reduced in these same genomic regions, thus eliminating explanations that rely on a correlation between the rates of mutation and crossing over. Several theoretical models (directional hitchhiking, background selection, and random environment) have been proposed as population genetic explanations. In this study samples from an African population (n = 50) and a European population (n = 51) were surveyed at the su(s) (1955 bp) and su(w(a)) (3213 bp) loci for DNA sequence polymorphism, utilizing a stratified SSCP/DNA sequencing protocol. These loci are located near the telomere of the X chromosome, in a region of reduced crossing over per physical length, and exhibit a significant reduction in DNA sequence polymorphism. Unlike most previously surveyed, these loci reveal substantial skews toward rare site frequencies, consistent with the predictions of directional hitchhiking and random environment models and inconsistent with the general predictions of the background selection model (or neutral theory). No evidence for excess geographic differentiation at these loci is observed. Although linkage disequilibrium is observed between closely linked sites within these loci, many recombination events in the genealogy of the sampled alleles can be inferred and the genomic scale of linkage disequilibrium, measured in base pairs between sites, is the same as that observed for loci in regions of normal crossing over. We conclude that gene conversion must be high in these regions of low crossing over.     

Background Selection as Baseline for Nucleotide Variation across the Drosophila Genome

The constant removal of deleterious mutations by natural selection causes a reduction in neutral diversity and efficacy of selection at genetically linked sites (a process called Background Selection, BGS). Population genetic studies, however, often ignore BGS effects when investigating demographic events or the presence of other types of selection. To obtain a more realistic evolutionary expectation that incorporates the unavoidable consequences of deleterious mutations, we generated high-resolution landscapes of variation across the Drosophila melanogaster genome under a BGS scenario independent of polymorphism data. We find that BGS plays a significant role in shaping levels of variation across the entire genome, including long introns and intergenic regions distant from annotated genes. We also find that a very large percentage of the observed variation in diversity across autosomes can be explained by BGS alone, up to 70% across individual chromosome arms at 100-kb scale, thus indicating that BGS predictions can be used as baseline to infer additional types of selection and demographic events. This approach allows detecting several outlier regions with signal of recent adaptive events and selective sweeps. The use of a BGS baseline, however, is particularly appropriate to investigate the presence of balancing selection and our study exposes numerous genomic regions with the predicted signature of higher polymorphism than expected when a BGS context is taken into account. Importantly, we show that these conclusions are robust to the mutation and selection parameters of the BGS model. Finally, analyses of protein evolution together with previous comparisons of genetic maps between Drosophila species, suggest temporally variable recombination landscapes and, thus, local BGS effects that may differ between extant and past phases. Because genome-wide BGS and temporal changes in linkage effects can skew approaches to estimate demographic and selective events, future analyses should incorporate BGS predictions and capture local recombination variation across genomes and along lineages.     

Multilocus patterns of polymorphism and selection across the X chromosome of Caenorhabditis remanei

Natural selection and neutral processes such as demography, mutation, and gene conversion all contribute to patterns of polymorphism within genomes. Identifying the relative importance of these varied components in evolution provides the principal challenge for population genetics. To address this issue in the nematode Caenorhabditis remanei, I sampled nucleotide polymorphism at 40 loci across the X chromosome. The site-frequency spectrum for these loci provides no evidence for population size change, and one locus presents a candidate for linkage to a target of balancing selection. Selection for codon usage bias leads to the non-neutrality of synonymous sites, and despite its weak magnitude of effect (N(e)s approximately 0.1), is responsible for profound patterns of diversity and divergence in the C. remanei genome. Although gene conversion is evident for many loci, biased gene conversion is not identified as a significant evolutionary process in this sample. No consistent association is observed between synonymous-site diversity and linkage-disequilibrium-based estimators of the population recombination parameter, despite theoretical predictions about background selection or widespread genetic hitchhiking, but genetic map-based estimates of recombination are needed to rigorously test for a diversity-recombination relationship. Coalescent simulations also illustrate how a spurious correlation between diversity and linkage-disequilibrium-based estimators of recombination can occur, due in part to the presence of unbiased gene conversion. These results illustrate the influence that subtle natural selection can exert on polymorphism and divergence, in the form of codon usage bias, and demonstrate the potential of C. remanei for detecting natural selection from genomic scans of polymorphism.     

Evidence of Selection upon Genomic GC-Content in Bacteria

The genomic GC-content of bacteria varies dramatically, from less than 20% to more than 70%. This variation is generally ascribed to differences in the pattern of mutation between bacteria. Here we test this hypothesis by examining patterns of synonymous polymorphism using datasets from 149 bacterial species. We find a large excess of synonymous GC→AT mutations over AT→GC mutations segregating in all but the most AT-rich bacteria, across a broad range of phylogenetically diverse species. We show that the excess of GC→AT mutations is inconsistent with mutation bias, since it would imply that most GC-rich bacteria are declining in GC-content; such a pattern would be unsustainable. We also show that the patterns are probably not due to translational selection or biased gene conversion, because optimal codons tend to be AT-rich, and the excess of GC→AT SNPs is observed in datasets with no evidence of recombination. We therefore conclude that there is selection to increase synonymous GC-content in many species. Since synonymous GC-content is highly correlated to genomic GC-content, we further conclude that there is selection on genomic base composition in many bacteria.     

Natural selection in gene-dense regions shapes the genomic pattern of polymorphism in wild and domesticated rice

Levels of nucleotide variability are frequently positively correlated with recombination rate and negatively associated with gene density due to the effects of selection on linked variation. These relationships are determined by properties that frequently differ among species, including the mating system, and aspects of genome organization such as how genes are distributed along chromosomes. In rice, genes are found at highest density in regions with frequent crossing-over. This association between gene density and recombination rate provides an opportunity to evaluate the effects of selection in a genomic context that differs from other model organisms. Using single-nucleotide polymorphism data from Asian domesticated rice Oryza sativa ssp. japonica and ssp. indica and their progenitor species O. rufipogon, we observe a significant negative association between levels of polymorphism and both gene and coding site density, but either no association, or a negative correlation, between nucleotide variability and recombination rate. We establish that these patterns are unlikely to be explained by neutral mutation rate biases and demonstrate that a model of background selection with variable rates of deleterious mutation is sufficient to account for the gene density effect in O. rufipogon. In O. sativa ssp. japonica, we report a strong negative correlation between polymorphism and recombination rate and greater losses of variation during domestication in the euchromatic chromosome arms than heterochromatin. This is consistent with Hill-Robertson interference in low-recombination regions, which may limit the efficacy of selection for domestication traits. Our results suggest that the physical distribution of selected mutations is a primary factor that determines the genomic pattern of polymorphism in wild and domesticated rice species.     

Pervasive recombination and sympatric genome diversification driven by frequency-dependent selection in Borrelia burgdorferi, the Lyme disease bacterium

How genomic diversity within bacterial populations originates and is maintained in the presence of frequent recombination is a central problem in understanding bacterial evolution. Natural populations of Borrelia burgdorferi, the bacterial agent of Lyme disease, consist of diverse genomic groups co-infecting single individual vertebrate hosts and tick vectors. To understand mechanisms of sympatric genome differentiation in B. burgdorferi, we sequenced and compared 23 genomes representing major genomic groups in North America and Europe. Linkage analysis of >13,500 single-nucleotide polymorphisms revealed pervasive horizontal DNA exchanges. Although three times more frequent than point mutation, recombination is localized and weakly affects genome-wide linkage disequilibrium. We show by computer simulations that, while enhancing population fitness, recombination constrains neutral and adaptive divergence among sympatric genomes through periodic selective sweeps. In contrast, simulations of frequency-dependent selection with recombination produced the observed pattern of a large number of sympatric genomic groups associated with major sequence variations at the selected locus. We conclude that negative frequency-dependent selection targeting a small number of surface-antigen loci (ospC in particular) sufficiently explains the maintenance of sympatric genome diversity in B. burgdorferi without adaptive divergence. We suggest that pervasive recombination makes it less likely for local B. burgdorferi genomic groups to achieve host specialization. B. burgdorferi genomic groups in the northeastern United States are thus best viewed as constituting a single bacterial species, whose generalist nature is a key to its rapid spread and human virulence.     

Natural Selection Constrains Neutral Diversity across A Wide Range of Species

The neutral theory of molecular evolution predicts that the amount of neutral polymorphisms within a species will increase proportionally with the census population size (Nc). However, this prediction has not been borne out in practice: while the range of Nc spans many orders of magnitude, levels of genetic diversity within species fall in a comparatively narrow range. Although theoretical arguments have invoked the increased efficacy of natural selection in larger populations to explain this discrepancy, few direct empirical tests of this hypothesis have been conducted. In this work, we provide a direct test of this hypothesis using population genomic data from a wide range of taxonomically diverse species. To do this, we relied on the fact that the impact of natural selection on linked neutral diversity depends on the local recombinational environment. In regions of relatively low recombination, selected variants affect more neutral sites through linkage, and the resulting correlation between recombination and polymorphism allows a quantitative assessment of the magnitude of the impact of selection on linked neutral diversity. By comparing whole genome polymorphism data and genetic maps using a coalescent modeling framework, we estimate the degree to which natural selection reduces linked neutral diversity for 40 species of obligately sexual eukaryotes. We then show that the magnitude of the impact of natural selection is positively correlated with Nc, based on body size and species range as proxies for census population size. These results demonstrate that natural selection removes more variation at linked neutral sites in species with large Nc than those with small Nc and provides direct empirical evidence that natural selection constrains levels of neutral genetic diversity across many species. This implies that natural selection may provide an explanation for this longstanding paradox of population genetics.     

Recombination rate variation in mice from an isolated island

Recombination rate is a heritable trait that varies among individuals. Despite the major impact of recombination rate on patterns of genetic diversity and the efficacy of selection, natural variation in this phenotype remains poorly characterized. We present a comparison of genetic maps, sampling 1212 meioses, from a unique population of wild house mice (Mus musculus domesticus) that recently colonized remote Gough Island. Crosses to a mainland reference strain (WSB/EiJ) reveal pervasive variation in recombination rate among Gough Island mice, including subchromosomal intervals spanning up to 28% of the genome. In spite of this high level of polymorphism, the genomewide recombination rate does not significantly vary. In general, we find that recombination rate varies more when measured in smaller genomic intervals. Using the current standard genetic map of the laboratory mouse to polarize intervals with divergent recombination rates, we infer that the majority of evolutionary change occurred in one of the two tested lines of Gough Island mice. Our results confirm that natural populations harbour a high level of recombination rate polymorphism and highlight the disparities in recombination rate evolution across genomic scales.     

Sympatric natural Saccharomyces cerevisiae and S. paradoxus populations have different thermal growth profiles

Saccharomyces cerevisiae and its close congener S. paradoxus are typically indistinguishable by the phenotypic criteria of classical yeast taxonomy, but they are evolutionarily distinct as indicated by hybrid spore inviability and genomic sequence divergence. Previous work has shown that these two species coexist in oak-associated microhabitats at natural woodland sites in North America. Here, we show that sympatric populations of S. cerevisiae and S. paradoxus from a single natural site are phenotypically differentiated in their growth rate responses to temperature. Our main finding is that the S. cerevisiae population exhibits a markedly higher growth rate at 37 degrees C than the S. paradoxus population; we also find possible differences in growth rate between these populations at two lower temperatures. We discuss the implications of our results for the coexistence of these yeasts in natural environments, and we suggest that thermal growth response may be an evolutionarily labile feature of these organisms that could be analyzed using genomic approaches.     

A short history of recombination in yeast

Despite it being the darling of fungal genomics, we know little about either the ecology or reproductive biology of the budding yeast, Saccharomyces cerevisiae, in nature. A recent study by Ruderfer et al. estimated that the ancestors of three S. cerevisiae genomes outcrossed approximately once every 50,000 generations, confirming the view that outcrossing is infrequent in natural populations of S. cerevisiae. This study also inferred the genomic positions of past recombination events. By comparing past recombination events with present-day recombination rates, this study lays the groundwork for determining whether recombination has improved the long-term survival of descendant lineages by bringing together favorable alleles, a longstanding question in evolutionary genetics.     

A Gene with Specific and Global Effects on Recombination of Sequences from Tandemly Repeated Genes in Saccharomyces Cerevisiae

The preservation of sequence homogeneity and copy number of tandemly repeated genes may require specific mechanisms or regulation of recombination. We have identified mutations that specifically affect recombination among natural repetitions in the yeast Saccharomyces cerevisiae. The rrm3 mutation stimulates mitotic recombination in the naturally occurring tandem repeats of the rDNA and copper chelatin (CUP1) genes. This mutation does not affect recombination of several other types of repeated genes tested including Ty elements, mating type information and duplications created by transformation. In addition to stimulating exchange among the multiple CUP1 repeats at their natural chromosomal location, rrm3 also increases recombination of a duplication of CUP1 units present at his4. This suggests that the RRM3 gene may encode a sequence-specific factor that contributes to a global suppression of mitotic exchange in sequences that can be maintained as tandem arrays.     

A composite-likelihood approach for detecting directional selection from DNA sequence data

We present a novel composite-likelihood-ratio test (CLRT) for detecting genes and genomic regions that are subject to recurrent natural selection (either positive or negative). The method uses the likelihood functions of Hartl et al. (1994) for inference in a Wright-Fisher genic selection model and corrects for nonindependence among sites by application of coalescent simulations with recombination. Here, we (1) characterize the distribution of the CLRT statistic (Lambda) as a function of the population recombination rate (R=4Ner); (2) explore the effects of bias in estimation of R on the size (type I error) of the CLRT; (3) explore the robustness of the model to population growth, bottlenecks, and migration; (4) explore the power of the CLRT under varying levels of mutation, selection, and recombination; (5) explore the discriminatory power of the test in distinguishing negative selection from population growth; and (6) evaluate the performance of maximum composite-likelihood estimation (MCLE) of the selection coefficient. We find that the test has excellent power to detect weak negative selection and moderate power to detect positive selection. Moreover, the test is quite robust to bias in the estimate of local recombination rate, but not to certain demographic scenarios such as population growth or a recent bottleneck. Last, we demonstrate that the MCLE of the selection parameter has little bias for weak negative selection and has downward bias for positively selected mutations.     

Molecular population genetics and evolution of a prion-like protein in Saccharomyces cerevisiae

The prion-like behavior of Sup35p, the eRF3 homolog in the yeast Saccharomyces cerevisiae, mediates the activity of the cytoplasmic nonsense suppressor known as [PSI(+)]. Sup35p is divided into three regions of distinct function. The N-terminal and middle (M) regions are required for the induction and propagation of [PSI(+)] but are not necessary for translation termination or cell viability. The C-terminal region encompasses the termination function. The existence of the N-terminal region in SUP35 homologs of other fungi has led some to suggest that this region has an adaptive function separate from translation termination. To examine this hypothesis, we sequenced portions of SUP35 in 21 strains of S. cerevisiae, including 13 clinical isolates. We analyzed nucleotide polymorphism within this species and compared it to sequence divergence from a sister species, S. paradoxus. The N domain of Sup35p is highly conserved in amino acid sequence and is highly biased in codon usage toward preferred codons. Amino acid changes are under weak purifying selection based on a quantitative analysis of polymorphism and divergence. We also conclude that the clinical strains of S. cerevisiae are not recently derived and that outcrossing between strains in S. cerevisiae may be relatively rare in nature.     

Constraints on the origin and maintenance of genetic kin recognition

Kin-recognition mechanisms allow helping behaviors to be directed preferentially toward related individuals, and could be expected to evolve in many cases. However, genetic kin recognition requires a genetic polymorphism on which recognition is based, and kin discriminating behaviors will affect the evolution of such polymorphism. It is unclear whether genetic polymorphisms used in kin recognition should be maintained by extrinsic selection pressures or not, as opposite conclusions have been reached by analytical one-locus models and simulations exploring different population structures. We analyze a two-locus model in a spatially subdivided population following the island model of dispersal between demes of finite size. We find that in the absence of mutation, selection eliminates polymorphism in most cases, except with extreme spatial structure and low recombination. With mutation, the population may reach a stable limit cycle over which both loci are polymorphic; however, the average frequency of conditional helping can be high only under strong structure and low recombination. Finally, we review evidence for extrinsic selection maintaining polymorphism on which kin recognition is based.     

Characterization of the slow-growth phenotype of S. cerevisiae Whip/Mgs1 Sgs1 double deletion mutants

RecQ DNA helicases from many organisms have been indicated to function in the maintenance of genomic stability. In human cells, mutation in the WRN helicase, a RecQ-like DNA helicase, results in the Werner syndrome (WS), a genetic disorder characterized by genomic instability and premature ageing. Similarly, mutation in SGS1, the RECQ homologue in budding yeast, results in genomic instability and accelerated ageing. We previously demonstrated that mouse WRN interacts physically with a novel, highly conserved protein that we named WHIP, and that in budding yeast cells, simultaneous deletion of WHIP/MGS1 and SGS1 results in slow growth and shortened life span. Here we show by using genetic analysis in Saccharomyces cerevisiae that mgs1Delta sgs1Delta cells have increased rates of terminal G2/M arrest, and show elevated rates of spontaneous sister chromatid recombination (SCR) and rDNA array recombination. Finally, we report that complementation of the synthetic relationship between SGS1 and WHIP/MGS1 requires both the helicase and Top3-binding activities of Sgs1, as well as the ATPase activity of Mgs1. Our results suggest that Whip/Mgs1 is implicated in DNA metabolism, and is required for normal growth and cell cycle progression in the absence of Sgs1.