Geographic variation in adaptation at the molecular level: a case study of plant immunity genes

Natural selection imposed by interacting species frequently varies among geographic locations and can lead to local adaptation, where alternative phenotypes are found in different populations. Little is known, however, about whether geographically variable selection acting on traits that mediate species interactions is consistent or strong enough to influence patterns of nucleotide variation at individual loci. To investigate this question, we examined patterns of nucleotide diversity and population structure at 16 plant innate immunity genes, with putative functions in defending plants against pathogens or herbivores, from six populations of teosinte (Zea mays ssp. parviglumis). Specifically, we tested whether patterns of population structure and within-population diversity at immunity genes differed from patterns found at nonimmunity (reference) loci and from neutral expectations derived from coalescent simulations of structured populations. For the majority of genes, we detected no strong evidence of geographically variable selection. However, in the wound-induced serine protease inhibitor (wip1), which inhibits the hydrolysis of dietary proteins in insect herbivores, one population showed unusually high levels of genetic differentiation, very low levels of nucleotide polymorphism, and was fixed for a novel replacement substitution in the active site of the protein. Taken together, these data suggest that wip1 experienced a recent selective sweep in one geographic region; this pattern may reflect local adaptation or an ongoing species-wide sweep. Overall, our results indicate that a signature of local adaptation at the molecular level may be uncommon-particularly for traits that are under complex genetic control.     

Comparative evolutionary histories of chitinase genes in the Genus zea and Family poaceae

Patterns of DNA sequence diversity vary widely among genes encoding proteins that protect plants against pathogens and herbivores. Comparative studies may help determine whether these differences are due to the strength of selection acting on different types of defense, in different evolutionary lineages, or both. I analyzed sequence diversity at three chitinases, a well-studied component of defense, in two species of Zea and several Poaceae taxa. Although the Zea species are closely related and these genes code for proteins with similar biochemical function, patterns of diversity vary widely within and among species. Intraspecific diversity at chiB, chiI, and Z. mays ssp. parviglumis chiA are consistent with a neutral-equilibrium model whereas chiA had no segregating sites within Z. diploperennis--consistent with a recent and strong selective sweep. Codons identified as having diverged among Poaceae taxa in response to positive selection were significantly overrepresented among targets of selection in Arabis, suggesting common responses to selection in distantly related plant taxa. Divergence of the recent duplicates chiA and chiB is consistent with positive selection but relaxed constraint cannot be rejected. Weak evidence for adaptive divergence of these duplicated downstream components of defense contrasts with strong evidence for adaptive divergence of genes involved in pathogen recognition.     

The molecular evolution of terminal ear1, a regulatory gene in the genus Zea.

Nucleotide diversity in the terminal ear1 (te1) gene, a regulatory locus hypothesized to be involved in the morphological evolution of maize (Zea mays ssp. mays), was investigated for evidence of past selection. Nucleotide polymorphism in a 1.4-kb region of te1 was analyzed for a sample of 26 sequences isolated from 12 maize lines, five populations of the maize progenitor, Z. mays ssp. parviglumis, six other Zea populations, and two Tripsacum species. Although nucleotide diversity in te1 in maize is reduced relative to ssp. parviglumis, phylogenetic and statistical analyses of the pattern of polymorphism among these sequences provided no evidence of past selection, indicating that the region of the gene studied was probably not involved in maize evolution. The level of reduction in genetic diversity in te1 in maize relative to its progenitor is comparable to that found in previous reports for isozymes and other neutrally evolving maize genes and is consistent with a genome-wide reduction of genetic diversity resulting from a domestication bottleneck. An estimate of the age (1.2-1.4 million yr) of the maize gene pool based on te1 is roughly consistent with previous estimates based on other neutral genes, but may be biased by the apparently slow synonymous substitution rate at te1.     

Population genetics of duplicated disease-defense genes,hm1 and hm2, in maize (Zea mays ssp. mays L.) and its wild ancestor (Zea mays ssp. parviglumis)

Plant defense genes are subject to nonneutral evolutionary dynamics. Here we investigate the evolutionary dynamics of the duplicated defense genes hm1 and hm2 in maize and its wild ancestor Zea mays ssp. parviglumis. Both genes have been shown to confer resistance to the fungal pathogen Cochliobolus carbonum race 1, but the effectiveness of resistance differs between loci. The genes also display different population histories. The hm1 locus has the highest nucleotide diversity of any gene yet sampled in the wild ancestor of maize, and it contains a large number of indel polymorphisms. There is no evidence, however, that high diversity in hm1 is a product of nonneutral evolution. In contrast, hm2 has very low nucleotide diversity in the wild ancestor of maize. The distribution of hm2 polymorphic sites is consistent with nonneutral evolution, as indicated by Tajima's D and other neutrality tests. In addition, one hm2 haplotype is more frequent than expected under the equilibrium neutral model, suggesting hitchhiking selection. Both defense genes retain >80% of the level of genetic variation in maize relative to the wild ancestor, and this level is similar to other maize genes that were not subject to artificial selection during domestication.     

Population genetic evidence for rapid changes in intraspecific diversity and allelic cycling of a specialist defense gene in Zea

Two patterns of plant defense gene evolution are emerging from molecular population genetic surveys. One is that specialist defenses experience stronger selection than generalist defenses. The second is that specialist defenses are more likely to be subject to balancing selection, i.e., evolve in a manner consistent with balanced-polymorphism or trench-warfare models of host-parasite coevolution. Because most of the data of specialist defenses come from Arabidopsis thaliana, we examined the genetic diversity and evolutionary history of three defense genes in two outcrossing species, the autotetraploid Zea perennis and its most closely related extant relative the diploid Z. diploperennis. Intraspecific diversity at two generalist defenses, the protease inhibitors wip1 and mpi, were consistent with a neutral model. Like previously studied genes in these taxa, wip1 and mpi harbored similar levels of diversity in Z. diploperennis and Z. perennis. In contrast, the specialist defense hm2 showed strong although distinctly different departures from a neutral model in the two species. Z. diploperennis appears to have experienced a strong and recent selective sweep. Using a rejection-sampling coalescent method, we estimate the strength of selection on Z. diploperennis hm2 to be approximately 3.0%, which is approximately equal to the strength of selection on tb1 during maize domestication. Z. perennis hm2 harbors three highly diverged alleles, two of which are found at high frequency. The distinctly different patterns of diversity may be due to differences in the phase of host-parasite coevolutionary cycles, although higher hm2 diversity in Z. perennis may also reflect reduced efficacy of selection in the autotetraploid relative to its diploid relative.     

Impact of demography on linked selection in two outcrossing Brassicaceae species

Genetic diversity is shaped by mutation, genetic drift, gene flow, recombination, and selection. The dynamics and interactions of these forces shape genetic diversity across different parts of the genome, between populations and species. Here, we have studied the effects of linked selection on nucleotide diversity in outcrossing populations of two Brassicaceae species, Arabidopsis lyrata and Capsella grandiflora, with contrasting demographic history. In agreement with previous estimates, we found evidence for a modest population size expansion thousands of generations ago, as well as efficient purifying selection in C. grandiflora. In contrast, the A. lyrata population exhibited evidence for very recent strong population size decline and weaker efficacy of purifying selection. Using multiple regression analyses with recombination rate and other genomic covariates as explanatory variables, we can explain 47% of the variance in neutral diversity in the C. grandiflora population, while in the A. lyrata population, only 11% of the variance was explained by the model. Recombination rate had a significant positive effect on neutral diversity in both species, suggesting that selection at linked sites has an effect on patterns of neutral variation. In line with this finding, we also found reduced neutral diversity in the vicinity of genes in the C. grandiflora population. However, in A. lyrata no such reduction in diversity was evident, a finding that is consistent with expectations of the impact of a recent bottleneck on patterns of neutral diversity near genes. This study thus empirically demonstrates how differences in demographic history modulate the impact of selection at linked sites in natural populations.     

Selection versus demography: a multilocus investigation of the domestication process in maize

The domestication of maize (Zea mays ssp. mays) from its wild ancestor (Zea mays ssp. parviglumis) led to a loss of genetic diversity both through a population bottleneck and through directional selection at agronomically important genes. In order to discriminate between those effects and to investigate the nature of the domestication bottleneck, we analyzed nucleotide diversity data from 12 chromosome 1 loci in parviglumis. We found an average loss of nucleotide diversity of 38% across genes, but this average was skewed downward by four putatively selected loci (tb1, d8, ts2, and zagl1). To better understand the domestication process, we used the coalescent with recombination to simulate bottlenecks under various lengths and population sizes. For each locus, we determine the likelihood of the observed data using three summary statistics: the number of segregating sites, an estimate of the population recombination parameter, and Tajima's D. Based on the eight neutrally evolving loci, a model with a bottleneck had a significantly higher likelihood than a model without one. The four putatively selected loci had significantly different likelihood optimums than the neutral loci, and this approach confirmed that ts2 and d8 were selected either during domestication or breeding. Overall, the best-fitting models had a bottleneck in which the population size and the bottleneck duration had a ratio of approximately 4- to approximately 5; for example, if the initial domestication event occurred over a 500-year period, the population size was roughly 2,000 to 2,500 individuals. However, this range did vary with the summary statistic used to assess the fit of simulations to data. In this context, Tajima's D performed poorly as a goodness-of-fit statistic, probably because Z. mays ssp. parviglumis has a frequency spectrum that is significantly skewed toward low-frequency variants. Finally, we found that demography is unlikely to account for the previously observed positive correlation between nucleotide diversity and the population-recombination parameter in maize, leaving this observation difficult to interpret.     

Population structure and its effects on patterns of nucleotide polymorphism in teosinte (Zea mays ssp. parviglumis)

Surveys of nucleotide diversity in the wild ancestor of maize, Zea mays ssp. parviglumis, have revealed genomewide departures from the standard neutral equilibrium (NE) model. Here we investigate the degree to which population structure may account for the excess of rare polymorphisms frequently observed in species-wide samples. On the basis of sequence data from five nuclear and two chloroplast loci, we found significant population genetic structure among seven subpopulations from two geographic regions. Comparisons of estimates of population genetic parameters from species-wide samples and subpopulation-specific samples showed that population genetic subdivision influenced observed patterns of nucleotide polymorphism. In particular, Tajima's D was significantly higher (closer to zero) in subpopulation-specific samples relative to species-wide samples, and therefore more closely corresponded to NE expectations. In spite of these overall patterns, the extent to which levels and patterns of polymorphism within subpopulations differed from species-wide samples and NE expectations depended strongly on the geographic region (Jalisco vs. Balsas) from which subpopulations were sampled. This may be due to the demographic history of subpopulations in those regions. Overall, these results suggest that explicitly accounting for population structure may be important for studies examining the genetic basis of ecologically and agronomically important traits as well as for identifying loci that have been the targets of selection.     

Speciation and domestication in maize and its wild relatives: evidence from the globulin-1 gene.

The grass genus Zea contains the domesticate maize and several wild taxa indigenous to Central and South America. Here we study the genetic consequences of speciation and domestication in this group by sampling DNA sequences from four taxa-maize (Zea mays ssp. mays), its wild progenitor (Z. mays ssp. parviglumis), a more distant species within the genus (Z. luxurians), and a representative of the sister genus (Tripsacum dactyloides). We sampled a total of 26 sequences from the glb1 locus, which encodes a nonessential seed storage protein. Within the Zea taxa sampled, the progenitor to maize contains the most sequence diversity. Maize contains 60% of the level of genetic diversity of its progenitor, and Z. luxurians contains even less diversity (32% of the level of diversity of Z. mays ssp. parviglumis). Sequence variation within the glb1 locus is consistent with neutral evolution in all four taxa. The glb1 data were combined with adh1 data from a previous study to make inferences about the population genetic histories of these taxa. Comparisons of sequence data between the two morphologically similar wild Zea taxa indicate that the species diverged approximately 700, 000 years ago from a common ancestor of intermediate size to their present populations. Conversely, the domestication of maize was a recent event that could have been based on a very small number of founding individuals. Maize retained a substantial proportion of the genetic variation of its progenitor through this founder event, but diverged rapidly in morphology.     

Coalescent patterns for chromosomal inversions in divergent populations

Chromosomal inversions allow genetic divergence of locally adapted populations by reducing recombination between chromosomes with different arrangements. Divergence between populations (or hybridization between species) is expected to leave signatures in the neutral genetic diversity of the inverted region. Quantitative expectations for these patterns, however, have not been obtained. Here, we develop coalescent models of neutral sites linked to an inversion polymorphism in two locally adapted populations. We consider two scenarios of local adaptation: selection on the inversion breakpoints and selection on alleles inside the inversion. We find that ancient inversion polymorphisms cause genetic diversity to depart dramatically from neutral expectations. Other situations, however, lead to patterns that may be difficult to detect; important determinants are the age of the inversion and the rate of gene flux between arrangements. We also study inversions under genetic drift, finding that they produce patterns similar to locally adapted inversions of intermediate age. Our results are consistent with empirical observations, and provide the foundation for quantitative analyses of the roles that inversions have played in speciation.     

Genomic screening for artificial selection during domestication and improvement in maize

BACKGROUND: Artificial selection results in phenotypic evolution. Maize (Zea mays L. ssp. mays) was domesticated from its wild progenitor teosinte (Zea mays subspecies parviglumis) through a single domestication event in southern Mexico between 6000 and 9000 years ago. This domestication event resulted in the original maize landrace varieties. The landraces provided the genetic material for modern plant breeders to select improved varieties and inbred lines by enhancing traits controlling agricultural productivity and performance. Artificial selection during domestication and crop improvement involved selection of specific alleles at genes controlling key morphological and agronomic traits, resulting in reduced genetic diversity relative to unselected genes. SCOPE: This review is a summary of research on the identification and characterization by population genetics approaches of genes affected by artificial selection in maize. CONCLUSIONS: Analysis of DNA sequence diversity at a large number of genes in a sample of teosintes and maize inbred lines indicated that approx. 2 % of maize genes exhibit evidence of artificial selection. The remaining genes give evidence of a population bottleneck associated with domestication and crop improvement. In a second study to efficiently identify selected genes, the genes with zero sequence diversity in maize inbreds were chosen as potential targets of selection and sequenced in diverse maize landraces and teosintes, resulting in about half of candidate genes exhibiting evidence for artificial selection. Extended gene sequencing demonstrated a low false-positive rate in the approach. The selected genes have functions consistent with agronomic selection for plant growth, nutritional quality and maturity. Large-scale screening for artificial selection allows identification of genes of potential agronomic importance even when gene function and the phenotype of interest are unknown. These approaches should also be applicable to other domesticated species if specific demographic conditions during domestication exist.     

Molecular evolution of the wound-induced serine protease inhibitor wip1 in Zea and related genera

Plant defense mechanisms have been the subject of intensive investigation. However, little is known about their long-term evolutionary dynamics. We investigated the molecular diversity of a wound-induced serine protease inhibitor, wip1, in the genus Zea, as well as the divergence of wip1 among four genera, Zea, Tripsacum, Sorghum, and Oryza, in order to gain insight into the long-term evolution of plant defense. The specific objectives of this study were to determine (1) whether wip1 has a history of positive or balancing selection, as has been shown for genes involved in plant defense against pathogens, and (2) if the evolutionary histories of wip1 inhibitory loops, which come into closest contact with proteases, differ from the evolutionary history of other parts of this gene. The Zea polymorphism data are consistent with a neutral evolutionary history. In contrast, relative-rate tests suggest a nonneutral evolutionary history. This inconsistency may indicate that selection acting on wip1 is episodic or that wip1 evolves in response to selection favoring novel alleles. We also detected significant heterogeneity in the evolutionary rates of the two inhibitory loops of wip1-one inhibitory loop is highly conserved, whereas the second has diverged rapidly. Because these two inhibitory loops are predicted to have very similar biochemical functions, the significantly different evolutionary histories suggest that these loops have different ecological functions.     

Contrasting patterns of diversity and population differentiation at the innate immunity gene toll-like receptor 2 (TLR2) in two sympatric rodent species

Comparing patterns of diversity and divergence between populations at immune genes and neutral markers can give insights into the nature and geographic scale of parasite-mediated selection. To date, studies investigating such patterns of selection in vertebrates have primarily focused on the acquired branch of the immune system, whereas it remains largely unknown how parasite-mediated selection shapes innate immune genes both within and across vertebrate populations. Here, we present a study on the diversity and population differentiation at the innate immune gene Toll-like receptor 2 (TLR2) across nine populations of yellow-necked mice (Apodemus flavicollis) and bank voles (Myodes glareolus) in southern Sweden. In yellow-necked mice, TLR2 diversity was very low, as was TLR2 population differentiation compared to neutral loci. In contrast, several TLR2 haplotypes co-occurred at intermediate frequencies within and across bank vole populations, and pronounced isolation by distance between populations was observed. The diversity and differentiation at neutral loci was similar in the two species. These results indicate that parasite-mediated selection has been acting in dramatically different ways on a given immune gene in ecologically similar and sympatric species. Furthermore, the finding of TLR2 population differentiation at a small geographical scale in bank voles highlights that vertebrate innate immune defense may be evolutionarily more dynamic than has previously been appreciated.     

Sequence diversity in the tetraploid Zea perennis and the closely related diploid Z. diploperennis: insights from four nuclear loci

Polyploidy has been an extremely common phenomenon in the evolutionary history of angiosperms. Despite this there are few data available to evaluate the effects of polyploidy on genetic diversity and to compare the relative effects of drift and selection in polyploids and related diploids. We investigated DNA sequence diversity at four nuclear loci (adh1, glb1, c1, and waxy) from the tetraploid Zea perennis and the closely related diploid Z. diploperennis. Contrary to expectations, we detected no strong evidence for greater genetic diversity in the tetraploid, or for consistent differences in the effects of either drift or selection between the tetraploid and the diploid. Our failure to find greater genetic diversity in Z. perennis may result from its relatively recent origin or demographic factors associated with its origin. In addition to comparing genetic diversity in the two species, we constructed genealogies to infer the evolutionary origin of Z. perennis. Although these genealogies are equivocal regarding the mode of origin, several aspects of these genealogies support an autotetraploid origin. Consistent with previous molecular data the genealogies do not, however, support the division of Zea into two sections, the section Zea and the section Luxuriantes.     

A large-scale screen for artificial selection in maize identifies candidate agronomic loci for domestication and crop improvement

Maize (Zea mays subsp mays) was domesticated from teosinte (Z. mays subsp parviglumis) through a single domestication event in southern Mexico between 6000 and 9000 years ago. This domestication event resulted in the original maize landrace varieties, which were spread throughout the Americas by Native Americans and adapted to a wide range of environmental conditions. Starting with landraces, 20th century plant breeders selected inbred lines of maize for use in hybrid maize production. Both domestication and crop improvement involved selection of specific alleles at genes controlling key morphological and agronomic traits, resulting in reduced genetic diversity relative to unselected genes. Here, we sequenced 1095 maize genes from a sample of 14 inbred lines and chose 35 genes with zero sequence diversity as potential targets of selection. These 35 genes were then sequenced in a sample of diverse maize landraces and teosintes and tested for selection. Using two statistical tests, we identified eight candidate genes. Extended gene sequencing of these eight candidate loci confirmed that six were selected throughout the gene, and the remaining two exhibited evidence of selection in the 3' portion of each gene. The selected genes have functions consistent with agronomic selection for nutritional quality, maturity, and productivity. Our large-scale screen for artificial selection allows identification of genes of potential agronomic importance even when gene function and the phenotype of interest are unknown.     

Molecular evolution of FLORICAULA/LEAFY orthologs in the Andropogoneae (Poaceae)

Members of the grass family (Poaceae) exhibit a broad range of inflorescence structures and other morphologies, making the grasses an interesting model system for studying the evolution of development. Here we present an analysis of the molecular evolution of FLORICAULA/LEAFY-like genes, which are important developmental regulatory loci known to affect inflorescence development in a wide range of flowering plant species. We have focused on sequences from the Andropogoneae, a tribe within the grass family that includes maize (Zea mays ssp. mays) and Sorghum (Sorghum bicolor). The FLORICAULA/LEAFY gene phylogeny we generated largely agrees with previously published phylogenies for the Andropogoneae using other nuclear genes but is unique in that it includes both members of one of the many duplicate gene sets present in maize. The placement of these sequences in the phylogeny suggests that the duplication of the maize FLORICAULA/LEAFY orthologs, zfl1 and zfl2, is a consequence of a proposed tetraploidy event that occurred in the common ancestor of Zea and a closely related genus, Tripsacum. Our data are consistent with the hypothesis that the transcribed regions of the FLORICAULA/LEAFY-like genes in the Andropogoneae are functionally constrained at both nonsynonymous and synonymous sites and show no evidence of directional selection. We also examined conservation of short noncoding sequences in the first intron, which may play a role in gene regulation. Finally, we investigated the genetic diversity of one of the two maize FLORICAULA/LEAFY orthologs, zfl2, in maize and its wild ancestor, teosinte (Z. mays ssp. parviglumis), and found no evidence for selection pressure resulting from maize domestication within the zfl2-coding region.     

A multilocus sequence survey in Arabidopsis thaliana reveals a genome-wide departure from a neutral model of DNA sequence polymorphism

The simultaneous analysis of multiple genomic loci is a powerful approach to studying the effects of population history and natural selection on patterns of genetic variation of a species. By surveying nucleotide sequence polymorphism at 334 randomly distributed genomic regions in 12 accessions of Arabidopsis thaliana, we examined whether a standard neutral model of nucleotide sequence polymorphism is consistent with observed data. The average nucleotide diversity was 0.0071 for total sites and 0.0083 for silent sites. Although levels of diversity are variable among loci, no correlation with local recombination rate was observed, but polymorphism levels were correlated for physically linked loci (<250 kb). We found that observed distributions of Tajima's D- and D/D(min)- and of Fu and Li's D-, D*- and F-, F*-statistics differed significantly from the expected distributions under a standard neutral model due to an excess of rare polymorphisms and high variances. Observed and expected distributions of Fay and Wu's H were not different, suggesting that demographic processes and not selection at multiple loci are responsible for the deviation from a neutral model. Maximum-likelihood comparisons of alternative demographic models like logistic population growth, glacial refugia, or past bottlenecks did not produce parameter estimates that were more consistent with observed patterns. However, exclusion of highly polymorphic "outlier loci" resulted in a fit to the logistic growth model. Various tests of neutrality revealed a set of candidate loci that may evolve under selection.     

High segregation distortion in maize B73 x teosinte crosses

Two genetic linkage maps of cultivated maize inbred lines and teosinte species were constructed. One population comprised 81 F(2) individuals derived from a cross between maize inbred line B73 and Zea mays ssp parviglumis, while the second consisted of 63 backcross individuals from a cross of maize inbred line B73 with Z. mays ssp diploperennis. In the B73 x Z. mays ssp parviglumis F(2) population, 172 simple sequence repeat (SSR) markers were mapped to 10 chromosomes, which covered 2210.8 cM. In the B73 x Z. mays ssp diploperennis backcross population, 258 SSR markers were mapped to 10 chromosomes, covering 1357.7 cM. Comparison of the two maps revealed that the total map length of Z. mays ssp diploperennis covers 1357.7 cM, which is about 61.4% of that of Z. mays ssp parviglumis (2210.8 cM). Extensive segregation distortion regions were found on chromosomes 1, 2, 3, 5, 6, 7, and 10 in the B73 x Z. mays ssp parviglumis F(2) population and on chromosomes 1-5 and 8-10 in the B73 x Z. mays ssp parviglumis backcross population. Segregation distortion analysis confirmed that the segregation distortion ratio in the interspecific population B73 x Z. mays ssp diploperennis was higher than in B73 x Z. mays ssp parviglumis. We found that the recombination distances are highly variable in these genetic crosses between cultivated and wild species of maize.     

Megabase-scale inversion polymorphism in the wild ancestor of maize

Chromosomal inversions are thought to play a special role in local adaptation, through dramatic suppression of recombination, which favors the maintenance of locally adapted alleles. However, relatively few inversions have been characterized in population genomic data. On the basis of single-nucleotide polymorphism (SNP) genotyping across a large panel of Zea mays, we have identified an ～50-Mb region on the short arm of chromosome 1 where patterns of polymorphism are highly consistent with a polymorphic paracentric inversion that captures >700 genes. Comparison to other taxa in Zea and Tripsacum suggests that the derived, inverted state is present only in the wild Z. mays subspecies parviglumis and mexicana and is completely absent in domesticated maize. Patterns of polymorphism suggest that the inversion is ancient and geographically widespread in parviglumis. Cytological screens find little evidence for inversion loops, suggesting that inversion heterozygotes may suffer few crossover-induced fitness consequences. The inversion polymorphism shows evidence of adaptive evolution, including a strong altitudinal cline, a statistical association with environmental variables and phenotypic traits, and a skewed haplotype frequency spectrum for inverted alleles.