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Model systems in biology expand the research capacity of individuals and the community. Closely related to Arabidopsis, the genus Boechera has emerged as an important ecological model owing to the ability to integrate across molecular, functional, and eco-evolutionary approaches. Boechera species are broadly distributed in relatively undisturbed habitats predominantly in western North America and provide one of the few experimental systems for identification of ecologically important genes through genome-wide association studies and investigations of selection with plants in their native habitats. The ecologically, evolutionarily, and agriculturally important trait of apomixis (asexual reproduction via seeds) is common in the genus, and field experiments suggest that abiotic and biotic environments shape the evolution of sex. To date, population genetic studies have focused on the widespread species B. stricta, detailing population divergence and demographic history. Molecular and ecological studies show that balancing selection maintains genetic variation in ~10% of the genome, and ecological trade-offs contribute to complex trait variation for herbivore resistance, flowering phenology, and drought tolerance. Microbiome analyses have shown that host genotypes influence leaf and root microbiome composition, and the soil microbiome influences flowering phenology and natural selection. Furthermore, Boechera offers numerous opportunities for investigating biological responses to global change. In B. stricta, climate change has induced a shift of2 weeks in the timing of first flowering since the 1970s, altered patterns of natural selection, generated maladaptation in previously locally-adapted populations, and disrupted life history trade-offs. Here we review resources and results for this eco-evolutionary model system and discuss future research directions.

Balancing selection is frequently invoked as a mechanism that maintains variation within and across populations. However, there are few examples of balancing selection operating on loci underpinning complex traits, which frequently display high levels of variation. We investigated mechanisms that may maintain variation in a focal polymorphism—leaf chemical profiles of a perennial wildflower (Boechera stricta, Brassicaceae)—explicitly interrogating multiple ecological and genetic processes including spatial variation in selection, antagonistic pleiotropy, and frequency-dependent selection. A suite of common garden and greenhouse experiments showed that the alleles underlying variation in chemical profile have contrasting fitness effects across environments, implicating two ecological drivers of selection on chemical profile: herbivory and drought. Phenotype-environment associations and molecular genetic analyses revealed additional evidence of past selection by these drivers. Together, these data are consistent with balancing selection on chemical profile, likely caused by pleiotropic effects of secondary chemical biosynthesis genes on herbivore defense and drought response., Editor’s Summary: Evidence for balancing selection acting on loci that control complex traits is limited. Here, the authors show evidence for past selection on chemical profile in a perennial wildflower by two ecological drivers, herbivory and drought, consistent with balancing selection on this trait.

The strength of selection varies among populations and across the genome, but the determinants of efficacy of selection remain unclear. In this study, we used whole-genome sequencing data from 467 Boechera stricta accessions to quantify the strength of selection and characterize the pattern of local adaptation. We found low genetic diversity on 0-fold degenerate sites and conserved non-coding sites, indicating functional constraints on these regions. The estimated distribution of fitness effects and the proportion of fixed substitutions suggest relaxed negative and positive selection in B. stricta. Among the four population groups, the NOR and WES groups have smaller effective population size (Ne), higher proportions of effectively neutral sites, and lower rates of adaptive evolution compared with UTA and COL groups, reflecting the effect of Ne on the efficacy of natural selection. We also found weaker selection on GC-biased sites compared with GC-conservative (unbiased) sites, suggested that GC-biased gene conversion has affected the strength of selection in B. stricta. We found mixed evidence for the role of the recombination rate on the efficacy of selection. The positive and negative selection was stronger in high-recombination regions compared with low-recombination regions in COL but not in other groups. By scanning the genome, we found different subsets of selected genes suggesting differential adaptation among B. stricta groups. These results show that differences in effective population size, nucleotide composition, and recombination rate are important determinants of the efficacy of selection. This study enriches our understanding of the roles of natural selection and local adaptation in shaping genomic variation.

When pleiotropy is present, genetic correlations may constrain the evolution of ecologically important traits. We used a quantitative genetics approach to investigate constraints on the evolution of secondary metabolites in a wild mustard, Boechera stricta. Much of the genetic variation in chemical composition of glucosinolates in B. stricta is controlled by a single locus, BCMA1/3. In a large-scale common garden experiment under natural conditions, we quantified fitness and glucosinolate profile in two leaf types and in fruits. We estimated genetic variances and covariances (the G-matrix) and selection on chemical profile in each tissue. Chemical composition of defenses was strongly genetically correlated between tissues. We found antagonistic selection between defense composition in leaves and fruits: compounds that were favored in leaves were disadvantageous in fruits. The positive genetic correlations and antagonistic selection led to strong constraints on the evolution of defenses in leaves and fruits. In a hypothetical population with no genetic variation at BCMA1/3, we found no evidence for genetic constraints, indicating that pleiotropy affecting chemical profile in multiple tissues drives constraints on the evolution of secondary metabolites.

Differential local adaptation restricts gene flow between populations inhabiting distinct environments, resulting in isolation by adaptation. In addition to the statistical inferences of genotype–environment associations, an integrative approach is needed to investigate the effect of local adaptation on population divergence at the ecological, genetic and genomic scale. Here, we combine reciprocal transplant, genome–environment association and QTL mapping to investigate local adaptation in Boechera stricta (Drummond's rockcress). With reciprocal transplant experiment, we found local genetic groups exhibit phenotypic characteristics corresponding to the distinct selection forces from different water availability. At the genetic level, the local allele of a major fitness QTL confers higher and sturdier flowering stalks, maximizing the fecundity fitness component under sufficient water supply, and its genetic variation is associated with precipitation across the landscape. At the genomewide scale, we further showed that multiple loci associated with precipitation are highly differentiated between genetic groups, suggesting that local adaptation has a widespread effect on reducing gene flow. This study provides one of the few comprehensive examples demonstrating how local adaptation facilitates population divergence at the trait, gene and genome level.

Differential local adaptation restricts gene flow between populations inhabiting distinct environments, resulting in isolation by adaptation. In addition to the statistical inferences of genotype–environment associations, an integrative approach is needed to investigate the effect of local adaptation on population divergence at the ecological, genetic and genomic scale. Here, we combine reciprocal transplant, genome–environment association and QTL mapping to investigate local adaptation in Boechera stricta (Drummond's rockcress). With reciprocal transplant experiment, we found local genetic groups exhibit phenotypic characteristics corresponding to the distinct selection forces from different water availability. At the genetic level, the local allele of a major fitness QTL confers higher and sturdier flowering stalks, maximizing the fecundity fitness component under sufficient water supply, and its genetic variation is associated with precipitation across the landscape. At the genomewide scale, we further showed that multiple loci associated with precipitation are highly differentiated between genetic groups, suggesting that local adaptation has a widespread effect on reducing gene flow. This study provides one of the few comprehensive examples demonstrating how local adaptation facilitates population divergence at the trait, gene and genome level.

Rice production is shifting from transplanting seedlings to direct sowing of seeds. Following heavy rains, directly sown seeds may need to germinate under anaerobic environments, but most rice (Oryza sativa) genotypes cannot survive these conditions. To identify the genetic architecture of complex traits, we quantified percentage anaerobic germination (AG) in 2,700 (wet-season) and 1,500 (dry-season) sequenced rice genotypes and performed genome-wide association studies (GWAS) using 693,502 single nucleotide polymorphisms. This was followed by post-GWAS analysis with a generalized SNP-to-gene set analysis, meta-analysis, and network analysis. We determined that percentage AG is intermediate-to-high among indica subpopulations, and AG is a polygenic trait associated with transcription factors linked to ethylene responses or genes involved in metabolic processes that are known to be associated with AG. Our post-GWAS analysis identified several genes involved in a wide variety of metabolic processes. We subsequently performed functional analysis focused on the small RNA and methylation pathways. We selected CLASSY 1 (CLSY1), a gene involved in the RNA-directed DNA methylation (RdDm) pathway, for further analyses under AG and found several lines of evidence that CLSY1 influences AG. We propose that the RdDm pathway plays a role in rice responses to water status during germination and seedling establishment developmental stages.

The genetic basis of flowering time changes across environments, and pleiotropy may limit adaptive evolution of populations in response to local conditions. However, little information is known about how genetic architecture changes among environments. We used genome-wide association studies (GWAS) in Boechera stricta (Graham) Al-Shehbaz, a relative of Arabidopsis, to examine flowering variation among environments and associations with climate conditions in home environments. Also, we used molecular population genetics to search for evidence of historical natural selection. GWAS found 47 significant quantitative trait loci (QTLs) that influence flowering time in one or more environments, control plastic changes in phenology between experiments, or show associations with climate in sites of origin. Genetic architecture of flowering varied substantially among environments. We found that some pairs of QTLs showed similar patterns of pleiotropy across environments. A large-effect QTL showed molecular signatures of adaptive evolution and is associated with climate in home environments. The derived allele at this locus causes later flowering and predominates in sites with greater water availability. This work shows that GWAS of climate associations and ecologically important traits across diverse environments can be combined with molecular signatures of natural selection to elucidate ecological genetics of adaptive evolution.

The mustard family (Brassicaceae) comprises several dozen monophyletic clades usually ranked as tribes. The tribe Boechereae plays a prominent role in plant research due to the incidence of apomixis and its close relationship to Arabidopsis. This tribe, largely confined to western North America, harbors nine genera and c. 130 species, with >90% of species belonging to the genus Boechera. Hundreds of apomictic diploid and triploid Boechera hybrids have spurred interest in this genus, but the remaining Boechereae genomes remain virtually unstudied. Here we report on comparative genome structure of six genera (Borodinia, Cusickiella, Phoenicaulis, Polyctenium, Nevada, and Sandbergia) and three Boechera species as revealed by comparative chromosome painting (CCP). All analyzed taxa shared the same seven-chromosome genome structure. Comparisons with the sister Halimolobeae tribe (n = 8) showed that the ancestral Boechereae genome (n = 7) was derived from an older n = 8 genome by descending dysploidy followed by the divergence of extant Boechereae taxa. As tribal divergence post-dated the origin of four tribe-specific chromosomes, it is proposed that these chromosomal rearrangements were a key evolutionary innovation underlaying the origin and diversification of the Boechereae in North America. Although most Boechereae genera exhibit genomic conservatism, intra-tribal cladogenesis has occasionally been accompanied by chromosomal rearrangements (particularly inversions). Recently, apomixis was reported in the Boechereae genera Borodinia and Phoenicaulis. Here, we report sexual reproduction in diploid Nevada, diploid Sandbergia, and tetraploid Cusickiella and aposporous apomixis in tetraploids of Polyctenium and Sandbergia. In sum, apomixis is now known to occur in five of the nine Boechereae genera.

Even after decades of research, the evolution of sex remains an enigma in evolutionary biology. Typically, research addresses the costs of sex and asexuality to characterize the circumstances in which one reproductive system is more favorable. Yet surprisingly few studies address the influence of common traits that are obligately correlated with asexuality, including hybridization and polyploidy; even though these traits have substantial impacts on selective patterns. In particular, hybridization is well-known to alter trait expression; these alterations may themselves represent a cost of sex. We examined the role of reproductive isolation in the formation of de novo hybrid lineages between two widespread species in the ecological model systemBoechera. Of 664 crosses betweenBoechera strictaandBoechera retrofracta, 17% of crosses produced F1 fruits. This suggests that postmating prezygotic barriers, i.e. pollen-pistil interactions, form the major barrier to hybrid success in this system. These interactions are asymmetrical, with 110 F1 fruits produced whenB. strictawas the maternal parent. This asymmetry was confirmed using a chloroplast phylogeny of wild-collectedB. stricta,B. retrofracta, and hybrids, which showed that most hybrids have aB. strictachloroplast haplotype. We next compared fitness of F2 hybrids and selfed parentalB. strictalines, finding that F2 fitness was reduced by substantial hybrid sterility. Our results suggest that multiple reproductively isolating barriers likely influence the formation and fitness of hybrid lineages in the wild, and that these costs of hybridization likely have profound impacts on the costs of sex in the natural environment.

In the natural world, sex prevails, despite its costs. Although much effort has been dedicated to identifying the intrinsic costs of sex (e.g., the cost of males), few studies have identified the ecological fitness consequences of sex. Furthermore, correlated biological traits that differ between sexuals and asexuals may alter these costs, or even render the typical costs of sex irrelevant. We conducted a large-scale, multisite, reciprocal transplant using multiple sexual and asexual genotypes of a native North American wildflower to show that sexual genotypes have reduced lifetime fitness, despite lower herbivory. We separated the effects of sex from those of hybridity, finding that overwinter survival is elevated in asexuals regardless of hybridity, but herbivores target hybrid asexuals more than nonhybrid asexual or sexual genotypes. Survival is lowest in homozygous sexual lineages, implicating inbreeding depression as a cost of sex. Our results show that the consequences of sex are shaped not just by sex itself, but by complex natural environments, correlated traits, and the identity and availability of mates.

The connections leading from genotype to fitness are not well understood, yet they are crucial for a diverse set of disciplines. Uncovering the general properties of biochemical pathways that influence ecologically important traits is an effective way to understand these connections. Enzyme flux control (or, control over pathway output) is one such pathway property. The flux-controlling enzyme in the antiherbivory aliphatic glucosinolate pathway of Arabidopsis thaliana has majority flux control under benign greenhouse conditions and has evidence of nonneutral evolution. However, it is unknown how patterns of flux control may change in different environments, or if insect herbivores respond to differences in pathway flux. We test this, first through genetic manipulation of the loci that code for the aliphatic glucosinolate pathway enzymes under a variety of environments (reduced water, reduced soil nutrients, leaf wounding and methyl jasmonate treatments), and find that flux control is consistently in the first enzyme of the pathway. We also find that a generalist herbivore, Trichoplusia ni, modifies its feeding behavior depending on the flux through the glucosinolate pathway. The influence over herbivore behavior combined with the consistency of flux control suggests that genes controlling flux might be repeatedly targeted by natural selection in diverse environments and species.

Freezing limits plant growth and crop productivity, and plant species in temperate zones have the capacity to develop freezing tolerance through complex modulation of gene expression affecting various aspects of metabolism and physiology. While many components of freezing tolerance have been identified in model species under controlled laboratory conditions, little is known about the mechanisms that impart freezing tolerance in natural populations of wild species. Here, we performed a quantitative trait locus (QTL) study of acclimated freezing tolerance in seedlings of Boechera stricta, a highly adapted relative of Arabidopsis (Arabidopsis thaliana) native to the Rocky Mountains. A single QTL was identified that contained the gene encoding ACYL-COENZYME A:DIACYLGLYCEROL ACYLTRANSFERASE1 (BstDGAT1), whose expression is highly cold responsive. The primary metabolic enzyme DGAT1 catalyzes the final step in assembly of triacylglycerol (TAG) by acyl transfer from acyl-CoA to diacylglycerol. Freezing tolerant plants showed higher DGAT1 expression during cold acclimation than more sensitive plants, and this resulted in increased accumulation of TAG in response to subsequent freezing. Levels of oligogalactolipids that are produced by SFR2 (SENSITIVE TO FREEZING2), an indispensable element of freezing tolerance in Arabidopsis, were also higher in freezing-tolerant plants. Furthermore, overexpression of AtDGAT1 led to increased freezing tolerance. We propose that DGAT1 confers freezing tolerance in plants by supporting SFR2-mediated remodeling of chloroplast membranes.

PREMISE OF THE STUDY: Ecological differentiation (ED) between sexual and asexual organisms may permit the maintenance of reproductive polymorphism. Several studies of sexual/asexual ED in plants have shown that the geographic ranges of asexuals extend beyond those of sexuals, often in areas of higher latitude or elevation. But very little is known about ED at fine scales, wherein coexistence of sexuals and asexuals may be permitted by differential niche occupation. METHODS: We used 149 populations of sexual and apomictic lineages in the genus Boechera (rock cress) collected across a portion of this mustard’s vast range. We characterized reproductive mode, ploidy, and species identity or hybrid parentage of each individual, and then used a multi-pronged statistical approach to 1) identify ED between sexuals and asexuals; 2) investigate the impacts of two confounding factors, polyploidy and hybridization, on ED; and 3) determine the environmental variables underlying ED. KEY RESULTS: We found that sexuals and asexuals are significantly ecologically differentiated across the landscape, despite fine-scale interdigitation of these two reproductive forms. Asexual reproduction was strongly associated with greater disturbance, reduced slope, and greater environmental variability. Although ploidy had little effect on the patterns observed, hybridization has a unique impact on the relationships between asexual reproduction and specific environmental variables. CONCLUSIONS: Ecological differentiation along the axes of disturbance, slope, and climatic variability, as well as the effects of heterozygosity, may contribute to the maintenance of sexuality and asexuality across the landscape, ultimately impacting the establishment and spread of asexual lineages.

Phenotypic plasticity is thought to impact evolutionary trajectories by shifting trait values in a direction that is either favored by natural selection (“adaptive plasticity”) or disfavored (“nonadaptive” plasticity). However, it is unclear how commonly each of these types of plasticity occurs in natural populations. To answer this question, we measured glucosinolate defensive chemistry and reproductive fitness in over 1,500 individuals of the wild perennial mustard Boechera stricta, planted in four common gardens across central Idaho, USA. Glucosinolate profiles—including total glucosinolate quantity as well as the relative abundances and overall diversity of different compounds—were strongly plastic both among habitats and within habitats. Patterns of glucosinolate plasticity varied greatly among genotypes. More often than expected by chance, glucosinolate profiles shifted in a direction that matched the direction of natural selection, indicating that plasticity among habitats tended to increase relative fitness. In contrast, we found no evidence for within-habitat selection on glucosinolate reaction norm slopes (i.e., plasticity along a continuous environmental gradient). Together, our results indicate that glucosinolate plasticity may improve the ability of B. stricta populations to persist after migration to new habitats.

Closely related to the model plant Arabidopsis thaliana, the genus Boechera is known to contain both sexual and apomictic species or accessions. Boechera retrofracta is a diploid sexually reproducing species and is thought to be an ancestral parent species of apomictic species. Here we report the de novo assembly of the B. retrofracta genome using short Illumina and Roche reads from 1 paired-end and 3 mate pair libraries. The distribution of 23-mers from the paired end library has indicated a low level of heterozygosity and the presence of detectable duplications and triplications. The genome size was estimated to be equal 227 Mb. N50 of the assembled scaffolds was 2.3 Mb. Using a hybrid approach that combines homology-based and de novo methods 27,048 protein-coding genes were predicted. Also repeats, transfer RNA (tRNA) and ribosomal RNA (rRNA) genes were annotated. Finally, genes of B. retrofracta and 6 other Brassicaceae species were used for phylogenetic tree reconstruction. In addition, we explored the histidine exonuclease APOLLO locus, related to apomixis in Boechera, and proposed model of its evolution through the series of duplications. An assembled genome of B. retrofracta will help in the challenging assembly of the highly heterozygous genomes of hybrid apomictic species.

Closely related to the model plant Arabidopsis thaliana, the genus Boechera is known to contain both sexual and apomictic species or accessions. Boechera retrofracta is a diploid sexually reproducing species and is thought to be an ancestral parent species of apomictic species. Here we report the de novo assembly of the B. retrofracta genome using short Illumina and Roche reads from 1 paired-end and 3 mate pair libraries. The distribution of 23-mers from the paired end library has indicated a low level of heterozygosity and the presence of detectable duplications and triplications. The genome size was estimated to be equal 227 Mb. N50 of the assembled scaffolds was 2.3 Mb. Using a hybrid approach that combines homology-based and de novo methods 27,048 protein-coding genes were predicted. Also repeats, transfer RNA (tRNA) and ribosomal RNA (rRNA) genes were annotated. Finally, genes of B. retrofracta and 6 other Brassicaceae species were used for phylogenetic tree reconstruction. In addition, we explored the histidine exonuclease APOLLO locus, related to apomixis in Boechera, and proposed model of its evolution through the series of duplications. An assembled genome of B. retrofracta will help in the challenging assembly of the highly heterozygous genomes of hybrid apomictic species.

Differences in tolerance to water stress may underlie ecological divergence of closely related ploidy lineages. However, the mechanistic basis of physiological variation governing ecogeographical cytotype segregation is not well understood. Here, using Brachypodium distachyon and its derived allotetraploid B. hybridum as model, we test the hypothesis that, for heteroploid annuals, ecological divergence of polyploids in drier environments is based on trait differentiation enabling drought escape. We demonstrate that under water limitation allotetraploids maintain higher photosynthesis and stomatal conductance and show earlier flowering than diploids, concordant with a drought-escape strategy to cope with water stress. Increased heterozygosity and greater genetic variability and plasticity of polyploids could confer a superior adaptive capability. Consistent with these predictions, we document (1) greater standing within-population genetic variation in water-use efficiency (WUE) and flowering time in allotetraploids, and (2) the existence of (nonlinear) environmental clines in physiology across allotetraploid populations. Increased gas exchange and diminished WUE occurred at the driest end of the gradient, consistent with a drought-escape strategy. Finally, we found that allotetraploids showed weaker genetic correlations than diploids congruous with the expectation of relaxed pleiotropic constraints in polyploids. Our results suggest evolutionary divergence of ecophysiological traits in each ploidy lineage.

Fixed chromosomal inversions can reduce gene flow and promote speciation in two ways: by suppressing recombination and by carrying locally favoured alleles at multiple loci. However, it is unknown whether favoured mutations slowly accumulate on older inversions or if young inversions spread because they capture pre-existing adaptive quantitative trait loci (QTLs). By genetic mapping, chromosome painting and genome sequencing, we have identified a major inversion controlling ecologically important traits in Boechera stricta. The inversion arose since the last glaciation and subsequently reached local high frequency in a hybrid speciation zone. Furthermore, the inversion shows signs of positive directional selection. To test whether the inversion could have captured existing, linked QTLs, we crossed standard, collinear haplotypes from the hybrid zone and found multiple linked phenology QTLs within the inversion region. These findings provide the first direct evidence that linked, locally adapted QTLs may be captured by young inversions during incipient speciation.Chromosome inversions play an important role in local adaptation and speciation1,2, and selectively important inversions have been identified in many species3,4. Selection due to different environmental factors or stages in the life cycle1 may favour inversions carrying locally adapted alleles at several loci. In addition, established inversions are predicted to accumulate selectively important genetic differences, which may contribute to reproductive isolation during speciation1.Although few studies have identified the actual loci that influence selection on inversions2,4,5, rearrangements may be favoured due to gene alterations near breakpoints6, chromatin changes7 or combinations of advantageous, co-adapted alleles8. Inversions suppress recombination, so locally advantageous alleles may segregate together, causing higher fitness than recombinant haplotypes9. Most evolutionary studies have focused on widespread, older inversions, so we have little knowledge of the evolutionary processes that guide their initial increase in frequency. For example, do inversions drift to higher frequency, and then acquire new advantageous mutations after they are common? Or are multiple linked, advantageous alleles captured in a new inversion, allowing them to spread together? Analysis of younger inversions may elucidate the evolutionary forces controlling the initial spread of chromosome inversions, which therefore influence their role in adaptation and speciation4,8.Related species often differ for chromosome inversions that carry locally favoured alleles at multiple loci10,11. A key distinction among models for the evolution of inversions is whether early frequency increase is due to genetic drift or natural selection. Genetic drift might predominate initially, with subsequent accumulation of advantageous variants12. Alternatively, the Kirkpatrick–Barton model9 argues that linked, locally adapted alleles exist first, and subsequently are captured within a new, selectively favoured inversion13. In this ‘inversion-late’ evolutionary sequence1,5, linked quantitative trait loci (QTLs), similar to the ancestral haplotype that gave rise to the inversion, may still exist in non-inverted genotypes9. Here, we test these predictions of the Kirkpatrick–Barton model. First, we introduce ecologically diverged subspecies of Boechera stricta. Next, we examine a young inversion to infer the selective forces controlling its early increase in frequency. Finally, we cross collinear, standard genotypes from the hybrid zone to ask whether old, linked QTLs can be found within the inversion region.

Many biological species are threatened with extinction because of a number of factors such as climate change and habitat loss, and their preservation depends on an accurate understanding of the extent of their genetic variability within and among populations. In this study, we assessed the genetic divergence of five quantitative traits in 10 populations of an endangered cruciferous species, Boechera fecunda, found in only several populations in each of two geographic regions (WEST and EAST) in southwestern Montana. We analyzed variation in quantitative traits, neutral molecular markers, and environmental factors and provided evidence that despite the restricted geographical distribution of this species, it exhibits a high level of genetic variation and regional adaptation. Conservation efforts therefore should be directed to the preservation of populations in each of these two regions without attempting transplantation between regions. Heritabilities and genetic coefficients of variation estimated from nested ANOVAs were generally high for leaf and rosette traits, although lower (and not significantly different from 0) for water-use efficiency. Measures of quantitative genetic differentiation, Q ST, were calculated for each trait from each pair of populations. For three of the five traits, these values were significantly higher between regions compared with those within regions (after adjustment for neutral genetic variation, F ST). This suggested that natural selection has played an important role in producing regional divergence in this species. Our analysis also revealed that the B. fecunda populations appear to be locally adapted due, at least in part, to differences in environmental conditions in the EAST and WEST regions.

Development of chilling and freezing tolerance is complex and can be affected by photoperiod, temperature and photosynthetic performance; however, there has been limited research on the interaction of these three factors. We evaluated 108 recombinant inbred lines of Boechera stricta, derived from a cross between lines originating from Montana and Colorado, under controlled long day (LD), short-day (SD) and in an outdoor environment (OE). We measured maximum quantum yield of photosystem II, lethal temperature for 50% survival and electrolyte leakage of leaves. Our results revealed significant variation for chilling and freezing tolerance and photosynthetic performance in different environments. Using both single- and multi-trait analyses, three main-effect quantitative trait loci (QTL) were identified. QTL on linkage group (LG)3 were SD specific, whereas QTL on LG4 were found under both LD and SD. Under all conditions, QTL on LG7 were identified, but were particularly predictive for the outdoor experiment. The co-localization of photosynthetic performance and freezing tolerance effects supports these traits being co-regulated. Finally, the major QTL on LG7 is syntenic to the Arabidopsis C-repeat binding factor locus, known regulators of chilling and freezing responses in Arabidopsis thaliana and other species.

Keywords: Flowering time; life history; microbiome; phenology; plant–microbe interactions; plasticity; selection; selective agents; soil ecology

The interactions between plants and enemies employ a variety of mechanisms, which in turn affect the long-term evolutionary histories of the interacting species. Different patterns of interactions determine not only the selective forces acting on individual genes, but also the flexibility and rapidity of evolution in response to new threats. New genomic techniques allowing large-scale comparisons within and between species demonstrate that defence genes are highly variable and experience very different selective patterns even in recently diverged species. Similar and even shared pathways for defence against diverse taxa of enemies reveal the variability and flexibility of plant defence.

Fitness trade-offs across episodes of selection and environments influence life-history evolution and adaptive population divergence. Documenting these trade-offs remains challenging as selection can vary in magnitude and direction through time and space. Here, we evaluate fitness trade-offs at the levels of the whole organism and the quantitative trait locus (QTL) in a multiyear field study of Boechera stricta (Brassicaceae), a genetically tractable mustard native to the Rocky Mountains. Reciprocal local adaptation was pronounced for viability, but not for reproductive components of fitness. Instead, local genomes had a fecundity advantage only in the high latitude garden. By estimating realized selection coefficients from individual-level data on viability and reproductive success and permuting the data to infer significance, we examined the genetic basis of fitness trade-offs. This analytical approach (Conditional Neutrality-Antagonistic Pleiotropy, CNAP) identified genetic trade-offs at a flowering phenology QTL (costs of adaptation) and revealed genetic trade-offs across fitness components (costs of reproduction). These patterns would not have emerged from traditional ANOVA-based QTL mapping. Our analytical framework can be applied to other systems to investigate fitness trade-offs. This task is becoming increasingly important as climate change may alter fitness landscapes, potentially disrupting fitness trade-offs that took many generations to evolve.

Ecological factors may contribute to reproductive isolation if differential local adaptation causes immigrant or hybrid fitness reduction. Since local adaptation results from the interaction between natural selection and adaptive traits, it is crucial to investigate both in order to understand ecological speciation. Previously, we used niche modeling to identify local water availability as an environmental correlate of incipient ecological speciation between two subspecies in Boechera stricta, a close relative of Arabidopsis. Here, we performed several large-scale greenhouse experiments to investigate the divergence of various physiological, phenological, and morphological traits. Although we found no significant difference in physiological traits, the Western subspecies has significantly faster growth rate, larger leaf area, less succulent leaves, delayed reproductive time, and longer flowering duration. These trait differences are concordant with previous results that habitats of the Western genotypes have more consistent water availability, while Eastern genotypes inhabit locations with more ephemeral water supplies. In addition, by comparing univariate and multivariate divergence of complex traits (QST) to the genome-wide distribution of SNP FST, we conclude that aspects of phenology and morphology (but not physiology) are under divergent selection. In addition, we also identified several highly diverged traits without obvious water-related functions.

The genetic mechanisms causing seed development by gametophytic apomixis in plants are predominantly unknown. As apomixis is consistently associated with hybridity and polyploidy, these confounding factors may either (a) be the underlying mechanism for the expression of apomixis, or (b) obscure the genetic factors which cause apomixis. To distinguish between these hypotheses, we analyzed the population genetic patterns of diploid and triploid apomictic lineages and their sexual progenitors in the genus Boechera (Brassicaceae). We find that while triploid apomixis is associated with hybridization, the majority of diploid apomictic lineages are likely the product of intra-specific crosses. We then show that these diploid apomicts are more likely to sire triploid apomictic lineages than conspecific sexuals. Combined with flow cytometric seed screen phenotyping for male and female components of apomixis, our analyses demonstrate that hybridization is an indirect correlate of apomixis in Boechera.

Adaptive evolution is shaped by the interaction of population genetics, natural selection and underlying network and biochemical constraints. Variation created by mutation, the raw material for evolutionary change, is translated into phenotypes by flux through metabolic pathways and by the topography and dynamics of molecular networks. Finally, the retention of genetic variation and the efficacy of selection depend on population genetics and demographic history. Emergent high-throughput experimental methods and sequencing technologies allow us to gather more evidence and to move beyond the theory in different systems and populations. Here we review the extent to which recent evidence supports long-established theoretical principles of adaptation.

Bacteria living on and in leaves and roots influence many aspects of plant health, so the extent of a plant's genetic control over its microbiota is of great interest to crop breeders and evolutionary biologists. Laboratory-based studies, because they poorly simulate true environmental heterogeneity, may misestimate or totally miss the influence of certain host genes on the microbiome. Here we report a large-scale field experiment to disentangle the effects of genotype, environment, age and year of harvest on bacterial communities associated with leaves and roots of Boechera stricta (Brassicaceae), a perennial wild mustard. Host genetic control of the microbiome is evident in leaves but not roots, and varies substantially among sites. Microbiome composition also shifts as plants age. Furthermore, a large proportion of leaf bacterial groups are shared with roots, suggesting inoculation from soil. Our results demonstrate how genotype-by-environment interactions contribute to the complexity of microbiome assembly in natural environments., Laboratory-based studies of the effect of plant genotype on plant microbiome composition often do not consider the influence of environmental heterogeneity. Here, Wagner et al. use a large-scale field experiment to assess the extent of host genetic and environmental factors on the microbiome of Boechera stricta.

Strategies that enable species to persist in changing environments have historically been divided into ecological (distributional shifts and phenotypic plasticity) and evolutionary (adaptation and gene flow). However, most species will likely need to rely on a combination of approaches to mitigate

Natural Selection at Work Catching the evolution of a novel function and determining its selective parameters in nature remains an extremely difficult task. Prasad et al. (p. 1081 ) have undertaken this quest documenting the molecular basis of a natural allelic polymorphism and its effects on herbivory and survival in the Arabidopsis relative, Boechera stricta , living in the Rocky Mountains.

Anthropogenic climate change has already altered the timing of major life-history transitions, such as the initiation of reproduction. Both phenotypic plasticity and adaptive evolution can underlie rapid phenological shifts in response to climate change, but their relative contributions are poorly understood. Here, we combine a continuous 38 year field survey with quantitative genetic field experiments to assess adaptation in the context of climate change. We focused on Boechera stricta (Brassicaeae), a mustard native to the US Rocky Mountains. Flowering phenology advanced significantly from 1973 to 2011, and was strongly associated with warmer temperatures and earlier snowmelt dates. Strong directional selection favoured earlier flowering in contemporary environments (2010–2011). Climate change could drive this directional selection, and promote even earlier flowering as temperatures continue to increase. Our quantitative genetic analyses predict a response to selection of 0.2 to 0.5 days acceleration in flowering per generation, which could account for more than 20 per cent of the phenological change observed in the long-term dataset. However, the strength of directional selection and the predicted evolutionary response are likely much greater now than even 30 years ago because of rapidly changing climatic conditions. We predict that adaptation will likely be necessary for long-term in situ persistence in the context of climate change.