The Roles of Drift and Selection on Short Stamen Loss in Arabidopsis thaliana along an Elevational Gradient in the Spanish Pyrenees.

Traits that have lost function sometimes persist through evolutionary time. These traits may be maintained by a lack of standing genetic variation for the trait, if selection against the trait is weak relative to drift, or if they have a residual function. To determine the evolutionary processes shaping whether nonfunctional traits are retained or lost, we investigated short stamens in 16 populations of Arabidopsis thaliana along an elevational cline in the Spanish Pyrenees. We found a cline in short stamen number from retention of short stamens in high elevation populations to incomplete loss in low elevation populations. We did not find evidence that limited genetic variation constrains the loss of short stamens at high elevations nor evidence for divergent selection on short stamens between high and low elevations. Finally, we identified loci associated with short stamens in the Spanish Pyrenees that are different from loci associated with variation in short stamen number across latitudes from a previous study. Overall, we did not identify the evolutionary mechanisms maintaining an elevational cline in short stamen number but did identify different genetic loci underlying the variation in short stamen along similar phenotypic clines.

A Multigenerational Turing Model Reproduces Transgressive Petal Spot Phenotypes in Hybrid Mimulus.

The origin of phenotypic novelty is a perennial question of genetics and evolution. To date, few studies of biological pattern formation specifically address multi-generational aspects of inheritance and phenotypic novelty. For quantitative traits influenced by many segregating alleles, offspring phenotypes are often intermediate to parental values. In other cases, offspring phenotypes can be transgressive to parental values. For example, in the model organism Mimulus (monkeyflower), the offspring of parents with solid-colored petals exhibit novel spotted petal phenotypes. These patterns are controlled by an activator-inhibitor gene regulatory network with a small number of loci. Here we develop and analyze a model of hybridization and pattern formation that accounts for the inheritance of a diploid gene regulatory network composed of either homozygous or heterozygous alleles. We find that the resulting model of multi-generational Turing-type pattern formation can reproduce transgressive petal phenotypes similar to those observed in Mimulus. The model gives insight into how non-patterned parent phenotypes can yield phenotypically transgressive, patterned offspring, aiding in the development of empirically testable hypotheses.

Temporal matches between monarch butterfly and milkweed population changes over the past 25,000 years.

In intimate ecological interactions, the interdependency of species may result in correlated demographic histories. For species of conservation concern, understanding the long-term dynamics of such interactions may shed light on the drivers of population decline. Here, we address the demographic history of the monarch butterfly, Danaus plexippus, and its dominant host plant, the common milkweed Asclepias syriaca (A. syriaca), using broad-scale sampling and genomic inference. Because genetic resources for milkweed have lagged behind those for monarchs, we first release a chromosome-level genome assembly and annotation for common milkweed. Next, we show that despite its enormous geographic range across eastern North America, A. syriaca is best characterized as a single, roughly panmictic population. Using approximate Bayesian computation with random forests (ABC-RF), a machine learning method for reconstructing demographic histories, we show that both monarchs and milkweed experienced population expansion during the most recent recession of North American glaciers 10,000-20,000 years ago. Our data also identify concurrent population expansions in both species during the large-scale clearing of eastern forests (∼200 years ago). Finally, we find no evidence that either species experienced a reduction in effective population size over the past 75 years. Thus, the well-documented decline of monarch abundance over the past 40 years is not visible in our genomic dataset, reflecting a possible mismatch of the overwintering census population to effective population size in this species.

Population genetics of transposable element load: A mechanistic account of observed overdispersion.

In an empirical analysis of transposable element (TE) abundance within natural populations of Mimulus guttatus and Drosophila melanogaster, we found a surprisingly high variance of TE count (e.g., variance-to-mean ratio on the order of 10 to 300). To obtain insight regarding the evolutionary genetic mechanisms that underlie the overdispersed population distributions of TE abundance, we developed a mathematical model of TE population genetics that includes the dynamics of element proliferation and purifying selection on TE load. The modeling approach begins with a master equation for a birth-death process and extends the predictions of the classical theory of TE dynamics in several ways. In particular, moment-based analyses of population distributions of TE load reveal that overdispersion is likely to arise via copy-and-paste proliferation dynamics, especially when the elementary processes of proliferation and excision are approximately balanced. Parameter studies and analytic work confirm this result and further suggest that overdispersed population distributions of TE abundance are probably not a consequence of purifying selection on total element load.

Range-wide variations in common milkweed traits and their effect on monarch larvae.

PREMISE: Leaf economic spectrum (LES) theory has historically been employed to inform vegetation models of ecosystem processes, but largely neglects intraspecific variation and biotic interactions. We attempt to integrate across environment-plant trait-herbivore interactions within a species at a range-wide scale. METHODS: We measured traits in 53 populations spanning the range of common milkweed (Asclepias syriaca) and used a common garden to determine the role of environment in driving patterns of intraspecific variation. We used a feeding trial to determine the role of plant traits in monarch (Danaus plexippus) larval development. RESULTS: Trait-trait relationships largely followed interspecific patterns in LES theory and persisted in a common garden when individual traits change. Common milkweed showed intraspecific variation and biogeographic clines in traits. Clines did not persist in a common garden. Larvae ate more and grew larger when fed plants with more nitrogen. A longitudinal environmental gradient in precipitation corresponded to a resource gradient in plant nitrogen, which produces a gradient in larval performance. CONCLUSIONS: Biogeographic patterns in common milkweed traits can sometimes be predicted from LES, are largely driven by environmental conditions, and have consequences for monarch larval performance. Changes to nutrient dynamics of landscapes with common milkweed could potentially influence monarch population dynamics. We show how biogeographic trends in intraspecific variation can influence key ecological interactions, especially in common species with large distributions.

Endosperm-based incompatibilities in hybrid monkeyflowers.

Endosperm is an angiosperm innovation central to their reproduction whose development, and thus seed viability, is controlled by genomic imprinting, where expression from certain genes is parent-specific. Unsuccessful imprinting has been linked to failed inter-specific and inter-ploidy hybridization. Despite their importance in plant speciation, the underlying mechanisms behind these endosperm-based barriers remain poorly understood. Here, we describe one such barrier between diploid Mimulus guttatus and tetraploid Mimulus luteus. The two parents differ in endosperm DNA methylation, expression dynamics, and imprinted genes. Hybrid seeds suffer from underdeveloped endosperm, reducing viability, or arrested endosperm and seed abortion when M. guttatus or M. luteus is seed parent, respectively, and transgressive methylation and expression patterns emerge. The two inherited M. luteus subgenomes, genetically distinct but epigenetically similar, are expressionally dominant over the M. guttatus genome in hybrid embryos and especially their endosperm, where paternal imprints are perturbed. In aborted seeds, de novo methylation is inhibited, potentially owing to incompatible paternal instructions of imbalanced dosage from M. guttatus imprints. We suggest that diverged epigenetic/regulatory landscapes between parental genomes induce epigenetic repatterning and global shifts in expression, which, in endosperm, may uniquely facilitate incompatible interactions between divergent imprinting schemes, potentially driving rapid barriers.

The regulatory network for petal anthocyanin pigmentation is shaped by the MYB5a/NEGAN transcription factor in Mimulus.

Much of the visual diversity of angiosperms is due to the frequent evolution of novel pigmentation patterns in flowers. The gene network responsible for anthocyanin pigmentation, in particular, has become a model for investigating how genetic changes give rise to phenotypic innovation. In the monkeyflower genus Mimulus, an evolutionarily recent gain of petal lobe anthocyanin pigmentation in M. luteus var. variegatus was previously mapped to genomic region pla2. Here, we use sequence and expression analysis, followed by transgenic manipulation of gene expression, to identify MYB5a-orthologous to the NEGAN transcriptional activator from M. lewisii-as the gene responsible for the transition to anthocyanin-pigmented petals in M. l. variegatus. In other monkeyflower taxa, MYB5a/NEGAN is part of a reaction-diffusion network that produces semi-repeating spotting patterns, such as the array of spots in the nectar guides of both M. lewisii and M. guttatus. Its co-option for the evolution of an apparently non-patterned trait-the solid petal lobe pigmentation of M. l. variegatus-illustrates how reaction-diffusion can contribute to evolutionary novelty in non-obvious ways. Transcriptome sequencing of a MYB5a RNAi line of M. l. variegatus reveals that this genetically simple change, which we hypothesize to be a regulatory mutation in cis to MYB5a, has cascading effects on gene expression, not only on the enzyme-encoding genes traditionally thought of as the targets of MYB5a but also on all of its known partners in the anthocyanin regulatory network.

Population genomic and historical analysis suggests a global invasion by bridgehead processes in Mimulus guttatus.

Imperfect historical records and complex demographic histories present challenges for reconstructing the history of biological invasions. Here, we combine historical records, extensive worldwide and genome-wide sampling, and demographic analyses to investigate the global invasion of Mimulus guttatus from North America to Europe and the Southwest Pacific. By sampling 521 plants from 158 native and introduced populations genotyped at >44,000 loci, we determined that invasive M. guttatus was first likely introduced to the British Isles from the Aleutian Islands (Alaska), followed by admixture from multiple parts of the native range. We hypothesise that populations in the British Isles then served as a bridgehead for vanguard invasions worldwide. Our results emphasise the highly admixed nature of introduced M. guttatus and demonstrate the potential of introduced populations to serve as sources of secondary admixture, producing novel hybrids. Unravelling the history of biological invasions provides a starting point to understand how invasive populations adapt to novel environments.

Author Correction: Origin and evolution of the octoploid strawberry genome.

In the version of this article originally published, author Joshua R. Puzey was incorrectly listed as having affiliation 7 (School of Plant Sciences, University of Arizona, Tucson, AZ, USA); affiliation 6 (Department of Biology, College of William and Mary, Williamsburg, VA, USA) is the correct affiliation. The error has been corrected in the HTML and PDF versions of the article.

A likelihood ratio test for changes in homeolog expression bias.

BACKGROUND: Gene duplications are a major source of raw material for evolution and a likely contributor to the diversity of life on earth. Duplicate genes (i.e., homeologs, in the case of a whole genome duplication) may retain their ancestral function, sub- or neofunctionalize, or be lost entirely. A primary way that duplicate genes evolve new functions is by altering their expression patterns. Comparing the expression patterns of duplicate genes gives clues as to whether any of these evolutionary processes have occurred. RESULTS: We develop a likelihood ratio test for the analysis of the expression ratios of duplicate genes across two conditions (e.g., tissues). We demonstrate an application of this test by comparing homeolog expression patterns of 1448 homeologous gene pairs using RNA-seq data generated from leaves and petals of an allotetraploid monkeyflower (Mimulus luteus). We assess the sensitivity of this test to different levels of homeolog expression bias and compare the method to several alternatives. CONCLUSIONS: The likelihood ratio test derived here is a direct, transparent, and easily implemented method for detecting changes in homeolog expression bias that outperforms alternative approaches. While our method was derived with homeolog analysis in mind, this method can be used to analyze changes in the ratio of expression levels between any two genes in any two conditions.

Origin and evolution of the octoploid strawberry genome.

Cultivated strawberry emerged from the hybridization of two wild octoploid species, both descendants from the merger of four diploid progenitor species into a single nucleus more than 1 million years ago. Here we report a near-complete chromosome-scale assembly for cultivated octoploid strawberry (Fragaria × ananassa) and uncovered the origin and evolutionary processes that shaped this complex allopolyploid. We identified the extant relatives of each diploid progenitor species and provide support for the North American origin of octoploid strawberry. We examined the dynamics among the four subgenomes in octoploid strawberry and uncovered the presence of a single dominant subgenome with significantly greater gene content, gene expression abundance, and biased exchanges between homoeologous chromosomes, as compared with the other subgenomes. Pathway analysis showed that certain metabolomic and disease-resistance traits are largely controlled by the dominant subgenome. These findings and the reference genome should serve as a powerful platform for future evolutionary studies and enable molecular breeding in strawberry.

Long- and short-term responses of Asclepias species differ in respect to fire, grazing, and nutrient addition.

PREMISE OF THE STUDY: The tallgrass prairie ecosystem has experienced a dramatic reduction over the past 150 yr. This reduction has impacted the abundance of native grassland species, including milkweeds (Asclepias). METHODS: We used two long-term (27 yr) data sets to examine how fire, grazing, and nutrient addition shape milkweed abundance in tallgrass prairie. We compared these results to those of a greenhouse experiment that varied nutrient levels in the absence of competition, herbivory, and mutualistic relationships. KEY RESULTS: Asclepias species exhibited broad patterns in response to burning regimes that did not include grazing, but experienced more species-specific patterns in other combinations. Asclepias syriaca was the only species to increase in abundance in plots that included burning and nutrient addition. In the greenhouse we found that nitrogen significantly increased biomass, while no effect of phosphorus was detected. CONCLUSIONS: These results indicate that A. syriaca will do best in settings with high nutrient loads, low competition, and no grazers. These characteristics define a small portion of the tallgrass prairie but exemplify modern agricultural settings, which have replaced prairies. However, other milkweeds examined did not share this pattern, which indicates that milkweed species will respond differently when exposed to agricultural settings, with some less able to cope with land conversion to pasture or row-crop agriculture.

Selective trade-offs maintain alleles underpinning complex trait variation in plants.

To understand evolutionary factors that maintain complex trait variation, we sequenced genomes from a single population of the plant Mimulus guttatus, identifying hundreds of nucleotide variants associated with morphological and life history traits. Alleles that delayed flowering also increased size at reproduction, which suggests pervasive antagonistic pleiotropy in this annual plant. The "large and slow" alleles, which were less common in small, rapidly flowering populations, became more abundant in populations with greater plant size. Furthermore, natural selection within the field population favored alternative alleles from year to year. Our results suggest that environmental fluctuations and selective trade-offs maintain polygenic trait variation within populations and also contribute to the geographic divergence in this wildflower species.

The causes and consequences of subgenome dominance in hybrids and recent polyploids.

Contents Summary 87 I. Introduction 87 II. Evolution in action: subgenome dominance within newly formed hybrids and polyploids 88 III. Summary and future directions 90 Acknowledgements 92 References 92 SUMMARY: The merger of divergent genomes, via hybridization or allopolyploidization, frequently results in a 'genomic shock' that induces a series of rapid genetic and epigenetic modifications as a result of conflicts between parental genomes. This conflict among the subgenomes routinely leads one subgenome to become dominant over the other subgenome(s), resulting in subgenome biases in gene content and expression. Recent advances in methods to analyze hybrid and polyploid genomes with comparisons to extant parental progenitors have allowed for major strides in understanding the mechanistic basis for subgenome dominance. In particular, our understanding of the role that homoeologous exchange might play in subgenome dominance and genome evolution is quickly growing. Here we describe recent discoveries uncovering the underlying mechanisms and provide a framework to predict subgenome dominance in hybrids and allopolyploids with far-reaching implications for agricultural, ecological, and evolutionary research.

Subgenome Dominance in an Interspecific Hybrid, Synthetic Allopolyploid, and a 140-Year-Old Naturally Established Neo-Allopolyploid Monkeyflower.

Recent studies have shown that one of the parental subgenomes in ancient polyploids is generally more dominant, having retained more genes and being more highly expressed, a phenomenon termed subgenome dominance. The genomic features that determine how quickly and which subgenome dominates within a newly formed polyploid remain poorly understood. To investigate the rate of emergence of subgenome dominance, we examined gene expression, gene methylation, and transposable element (TE) methylation in a natural, <140-year-old allopolyploid (Mimulus peregrinus), a resynthesized interspecies triploid hybrid (M. robertsii), a resynthesized allopolyploid (M. peregrinus), and progenitor species (M. guttatus and M. luteus). We show that subgenome expression dominance occurs instantly following the hybridization of divergent genomes and significantly increases over generations. Additionally, CHH methylation levels are reduced in regions near genes and within TEs in the first-generation hybrid, intermediate in the resynthesized allopolyploid, and are repatterned differently between the dominant and recessive subgenomes in the natural allopolyploid. Subgenome differences in levels of TE methylation mirror the increase in expression bias observed over the generations following hybridization. These findings provide important insights into genomic and epigenomic shock that occurs following hybridization and polyploid events and may also contribute to uncovering the mechanistic basis of heterosis and subgenome dominance.

A Whole-Transcriptome Approach to Evaluating Reference Genes for Quantitative Gene Expression Studies: A Case Study in Mimulus.

While quantitative PCR (qPCR) is widely recognized as being among the most accurate methods for quantifying gene expression, it is highly dependent on the use of reliable, stably expressed reference genes. With the increased availability of high-throughput methods for measuring gene expression, whole-transcriptome approaches may be increasingly utilized for reference gene selection and validation. In this study, RNA-seq was used to identify a set of novel qPCR reference genes and evaluate a panel of traditional "housekeeping" reference genes in two species of the evolutionary model plant genus Mimulus More broadly, the methods proposed in this study can be used to harness the power of transcriptomes to identify appropriate reference genes for qPCR in any study organism, including emerging and nonmodel systems. We find that RNA-seq accurately estimates gene expression means in comparison to qPCR, and that expression means are robust to moderate environmental and genetic variation. However, measures of expression variability were only in agreement with qPCR for samples obtained from a shared environment. This result, along with transcriptome-wide comparisons, suggests that environmental changes have greater impacts on expression variability than on expression means. We discuss how this issue can be addressed through experimental design, and suggest that the ever-expanding pool of published transcriptomes represents a rich and low-cost resource for developing better reference genes for qPCR.

Population structure and local selection yield high genomic variation in Mimulus guttatus.

Across western North America, Mimulus guttatus exists as many local populations adapted to site-specific environmental challenges. Gene flow between locally adapted populations will affect genetic diversity both within demes and across the larger metapopulation. Here, we analyse 34 whole-genome sequences from the intensively studied Iron Mountain population (IM) in conjunction with sequences from 22 Mimulus individuals sampled from across western North America. Three striking features of these data address hypotheses about migration and selection in a locally adapted population. First, we find very high levels of intrapopulation polymorphism (synonymous π = 0.033). Variation outside of genes is likely even higher but difficult to estimate because excessive divergence reduces the efficiency of read mapping. Second, IM exhibits a significantly positive genomewide average for Tajima's D. This indicates allele frequencies are typically more intermediate than expected from neutrality, opposite the pattern observed in many other species. Third, IM exhibits a distinctive haplotype structure with a genomewide excess of positive associations between rarer alleles at linked loci. This suggests an important effect of gene flow from other Mimulus populations, although a residual effect of population founding might also contribute. The combination of multiple analyses, including a novel tree-based analytic method, illustrates how the balance of local selection, limited dispersal and metapopulation dynamics manifests across the genome. The overall genomic pattern of sequence diversity suggests successful gene flow of divergent immigrant genotypes into IM. However, many loci show patterns indicative of local adaptation, particularly at SNPs associated with chromosomal inversions.

Geographic Variation of Plant Circadian Clock Function in Natural and Agricultural Settings.

The increasing demand for improved agricultural production will require more efficient breeding for traits that maintain yield under heterogeneous environments. The internal circadian oscillator is essential for perceiving and coordinating environmental cues such as day length, temperature, and abiotic stress responses within physiological processes. To investigate the contribution of the circadian clock to local adaptability, we have analyzed circadian period by leaf movement in natural populations of Mimulus guttatus and domesticated cultivars of Glycine max. We detected consistent variation in circadian period along a latitudinal gradient in annual populations of the wild plant and the selectively bred crop, and this provides novel evidence of natural and artificial selection for circadian performance. These findings provide new support that the circadian clock acts as a central regulator of plant adaptability and further highlight the potential of applying circadian clock gene variation to marker-assisted breeding programs in crops.

Strongly asymmetric hybridization barriers shape the origin of a new polyploid species and its hybrid ancestor.

PREMISE OF THE STUDY: Hybridization between diploids and tetraploids can lead to new allopolyploid species, often via a triploid intermediate. Viable triploids are often produced asymmetrically, with greater success observed for "maternal-excess" crosses where the mother has a higher ploidy than the father. Here we investigated the evolutionary origins of Mimulus peregrinus, an allohexaploid recently derived from the triploid M. ×robertsii, to determine whether reproductive asymmetry has shaped the formation of this new species. METHODS: We used reciprocal crosses between the diploid (M. guttatus) and tetraploid (M. luteus) progenitors to determine the viability of triploid M. ×robertsii hybrids resulting from paternal- vs. maternal-excess crosses. To investigate whether experimental results predict patterns seen in the field, we performed parentage analyses comparing natural populations of M. peregrinus to its diploid, tetraploid, and triploid progenitors. Organellar sequences obtained from pre-existing genomic data, supplemented with additional genotyping was used to establish the maternal ancestry of multiple M. peregrinus and M. ×robertsii populations. KEY RESULTS: We found strong evidence for asymmetric origins of M. peregrinus, but opposite to the common pattern, with paternal-excess crosses significantly more successful than maternal-excess crosses. These results successfully predicted hybrid formation in nature: 111 of 114 M. ×robertsii individuals, and 27 of 27 M. peregrinus, had an M. guttatus maternal haplotype. CONCLUSION: This study, which includes the first Mimulus chloroplast genome assembly, demonstrates the utility of parentage analysis through genome skimming. We highlight the benefits of complementing genomic analyses with experimental approaches to understand asymmetry in allopolyploid speciation.