Does selection occur at the intermediate zone of two insufficiently isolated populations? A whole-genome analysis along an altitudinal gradient.

Adaptive divergence occurs even between insufficiently isolated populations when there is a great difference in environments between their habitats. Individuals present in an intermediate zone of the two divergent populations are expected to have an admixed genetic structure due to gene flow. A selective pressure that acts on the genetically admixed individuals may limit the gene flow and maintain the adaptive divergence. Here, we addressed a question whether selection occurs in the genetically admixed individuals between two divergent populations. Arabidopsis halleri is a perennial montane plant, which has clear phenotypic dimorphisms between highland and lowland habitats in Mt. Ibuki, central Japan. We obtained the whole-genome sequences of Arabidopsis halleri plants along an altitudinal gradient of 359-1,317 m with a high spatial resolution (mean altitudinal interval of 20 m). We found a zone where the highland and lowland genes were mixing (intermediate subpopulation). In the intermediate subpopulation, we identified 5 and 13 genome regions, which included 3 and 8 genes, that had a high frequency of alleles that are accumulated in highland and lowland subpopulations, respectively. In addition, we also found that the frequency of highland alleles of these selected genome regions was smaller in the lowland subpopulation compared with that of the non-selected regions. These results suggest that the selection in the intermediate subpopulation might limit the gene flow and contribute to the adaptive divergence between altitudes. We also identified 7 genome regions that had low heterozygote frequencies in the intermediate subpopulation. We conclude that different types of selection in addition to gene flow occur at the intermediate altitude and shape the genetic structure across altitudes.

Molecular mechanism underlying pseudopeloria in Habenaria radiata (Orchidaceae).

Habenaria radiata (Orchidaceae) has two whorls of perianth, comprising three greenish sepals, two white petals and one lip (labellum). By contrast, the pseudopeloric (with a decreased degree of zygomorphy) mutant cultivar of H. radiata, 'Hishou', has changes in the identities of the dorsal sepal to a petaloid organ and the two ventral sepals to lip-like organs. Here, we isolated four DEFICIENS-like and two AGL6-like genes from H. radiata, and characterized their expression. Most of these genes revealed similar expression patterns in the wild type and in the 'Hishou' cultivar, except HrDEF-C3. The HrDEF-C3 gene was expressed in petals and lip in the wild type but was ectopically expressed in sepal, petals, lip, leaf, root and bulb in 'Hishou'. Sequence analysis of the HrDEF-C3 loci revealed that the 'Hishou' genome harbored two types of HrDEF-C3 genes: one identical to wild-type HrDEF-C3 and the other carrying a retrotransposon insertion in its promoter. Genetic linkage analysis of the progeny derived from an intraspecific cross between 'Hishou' and the wild type demonstrated that the mutant pseudopeloric trait was dominantly inherited and was linked to the HrDEF-C3 gene carrying the retrotransposon. These results indicate that the pseudopeloric phenotype is caused by retrotransposon insertion in the HrDEF-C3 promoter, resulting in the ectopic expression of HrDEF-C3. As the expression of HrAGL6-C2 was limited to lateral sepals and lip, the overlapping expression of HrDEF-C3 and HrAGL6-C2 is likely to be responsible for the sepal to lip-like identity in the lateral sepals of the 'Hishou' cultivar.

A Genome Scan for Genes Underlying Microgeographic-Scale Local Adaptation in a Wild Arabidopsis Species.

Adaptive divergence at the microgeographic scale has been generally disregarded because high gene flow is expected to disrupt local adaptation. Yet, growing number of studies reporting adaptive divergence at a small spatial scale highlight the importance of this process in evolutionary biology. To investigate the genetic basis of microgeographic local adaptation, we conducted a genome-wide scan among sets of continuously distributed populations of Arabidopsis halleri subsp. gemmifera that show altitudinal phenotypic divergence despite gene flow. Genomic comparisons were independently conducted in two distinct mountains where similar highland ecotypes are observed, presumably as a result of convergent evolution. Here, we established a de novo reference genome and employed an individual-based resequencing for a total of 56 individuals. Among 527,225 reliable SNP loci, we focused on those showing a unidirectional allele frequency shift across altitudes. Statistical tests on the screened genes showed that our microgeographic population genomic approach successfully retrieve genes with functional annotations that are in line with the known phenotypic and environmental differences between altitudes. Furthermore, comparison between the two distinct mountains enabled us to screen out those genes that are neutral or adaptive only in either mountain, and identify the genes involved in the convergent evolution. Our study demonstrates that the genomic comparison among a set of genetically connected populations, instead of the commonly-performed comparison between two isolated populations, can also offer an effective screening for the genetic basis of local adaptation.

Molecular phylogeny of the genus Asparagus (Asparagaceae) explains interspecific crossability between the garden asparagus (A. officinalis) and other Asparagus species.

The genus Asparagus comprises approximately 200 species, some of which are commercially cultivated, such as the garden asparagus (A. officinalis). Many Asparagus species, including A. officinalis, are dioecious and have been grouped into a subgenus distinct from that of hermaphroditic species. Although many interspecific crossings have been attempted to introduce useful traits into A. officinalis, only some of the dioecious species were found to be cross-compatible with A. officinalis. Here, molecular phylogenetic analyses were conducted to determine whether interspecific crossability is proportional to the genetic distance between the crossing pairs and to further clarify the evolutionary history of the Asparagus genus. A clade with all cross-compatible species and no cross-incompatible species was recovered in the phylogenetic tree based on analyses of non-coding cpDNA regions. In addition, a sex-linked marker developed for A. officinalis amplified a male-specific region in all cross-compatible species. The phylogenetic analyses also provided some insights about the evolutionary history of Asparagus; for example, by indicating that the genus had its origin in southern Africa, subsequently spreading throughout the old world through intensive speciation and dispersal. The results also suggest that dioecious species were derived from a single evolutionary transition from hermaphroditism in Asparagus. These findings not only contribute towards the understanding of the evolutionary history of the genus but may also facilitate future interspecific hybridization programs involving Asparagus species.

The evolution of self-compatible and self-incompatible populations in a hermaphroditic perennial, Trillium camschatcense (Melanthiaceae).

The evolution of selfing from outcrossing ancestors is known to have occurred repeatedly in angiosperms. Theoretical studies have argued that the transition from outcrossing to selfing is accomplished more easily than the reverse case, and phylogenetic analyses involving self-compatible (SC) and self-incompatible (SI) species has basically supported this assumption. The evolutionary direction of self-compatibility and self-incompatibility was examined in Trillium camschatcense, which contains geographically widespread SC populations, and restricted SI populations. Ecological surveys have revealed that the SC populations were suitable for outcrossing, and selfing in these populations did not confer any fitness advantage. Since reproductive fitness indicates the possibility of an evolutionary shift from self-compatibility to self-incompatibility, the phylogenetic relationships of SI and SC populations of T. camschatcense were investigated based on cpDNA variations and nuclear DNA microsatellite polymorphisms. Although phylogenetic analyses did not provide credible evidence to determine evolutionary direction, the SI populations turned out to be monophyletic with extremely low genetic differentiation. Based on these results, we proposed two possible scenarios for the evolutionary backgrounds of SI and SC populations in T. camschatcense. The plausibility of each scenario was evaluated based on the reproductive and geographical features of the mating systems.

Suppression of B function strongly supports the modified ABCE model in Tricyrtis sp. (Liliaceae).

B class MADS-box genes play important roles in petal and stamen development. Some monocotyledonous species, including liliaceous ones, produce flowers with petaloid tepals in whorls 1 and 2. A modified ABCE model has been proposed to explain the molecular mechanism of development of two-layered petaloid tepals. However, direct evidence for this modified ABCE model has not been reported to date. To clarify the molecular mechanism determining the organ identity of two-layered petaloid tepals, we used chimeric repressor gene-silencing technology (CRES-T) to examine the suppression of B function in the liliaceous ornamental Tricyrtis sp. Transgenic plants with suppressed B class genes produced sepaloid tepals in whorls 1 and 2 instead of the petaloid tepals as expected. In addition, the stamens of transgenic plants converted into pistil-like organs with ovule- and stigma-like structures. This report is the first to describe the successful suppression of B function in monocotyledonous species with two-layered petaloid tepals, and the results strongly support the modified ABCE model.

Analysis of the floral MADS-box genes from monocotyledonous Trilliaceae species indicates the involvement of SEPALLATA3-like genes in sepal-petal differentiation.

The evolution of greenish sepals from petaloid outer tepals has occurred repeatedly in various lineages of non-grass monocots. Studies in distinct monocot species showed that the evolution of sepals could be explained by the ABC model; for example, the defect of B-class function in the outermost whorl was linked to the evolution of sepals. Here, floral MADS-box genes from three sepal-bearing monocotyledonous Trilliaceae species, Trillium camschatcense, Paris verticillata, and Kinugasa japonica were examined. Unexpectedly, expression of not only A- but also B-class genes was detected in the sepals of all three species. Although the E-class gene is generally expressed across all floral whorls, no expression was detected in sepals in the three species examined here. Overexpression of the E-class SEPALLATA3-like gene from T. camschatcense (TcamSEP) in Arabidopsis thaliana produced phenotypes identical to those reported for orthologs in other monocots. Additionally, yeast hybrid experiments indicated that TcamSEP could form a higher-order complex with an endogenous heterodimer of B-class APETALA3/DEFICIENS-like (TcamDEF) and PISTILLATA/GLOBOSA-like (TcamGLO) proteins. These results suggest a conserved role for Trilliaceae SEPALLATA3-like genes in functionalization of the B-class genes, and that a lack of SEPALLATA3-like gene expression in the outermost whorl may be related to the formation of greenish sepals.

Adaptive significance of self-fertilization in a hermaphroditic perennial, Trillium camschatcense (Melanthiaceae).

The evolution of self-fertilization from primarily outcrossing ancestors is one of the most common evolutionary transitions in plants; however, the ecological mechanisms that maintain self-fertilization have remained controversial. Theoretical studies suggest that selfing is advantageous over outcrossing in terms of genetic transmission and assurance of seed production under pollen-limited circumstances. Trillium camschatcense is a herbaceous perennial distributed in Hokkaido and northern Honshu, Japan. Geographical variation in the breeding system (self-compatible, SC; or self-incompatible, SI) has been reported in populations in Hokkaido. Here, we used several SC and SI populations of T. camschatcense to investigate the adaptive significance and the evolutionary basis of self-fertilization. Pollination experiments and genetic analyses demonstrated that the potential availability of outcross pollen in SC populations was sufficient and that the number of pollen donors was equal to that of SI populations. However, despite the high availability of outcross pollen, the SC populations produced seeds predominantly by selfing and so underwent severe inbreeding depression. Although none of the suggested advantages for self-fertilization were supported by our analyses, we propose two possible scenarios for the evolution of self-fertilization in T. camschatcense.