Adaptive reduction of male gamete number in the selfing plant Arabidopsis thaliana.

The number of male gametes is critical for reproductive success and varies between and within species. The evolutionary reduction of the number of pollen grains encompassing the male gametes is widespread in selfing plants. Here, we employ genome-wide association study (GWAS) to identify underlying loci and to assess the molecular signatures of selection on pollen number-associated loci in the predominantly selfing plant Arabidopsis thaliana. Regions of strong association with pollen number are enriched for signatures of selection, indicating polygenic selection. We isolate the gene REDUCED POLLEN NUMBER1 (RDP1) at the locus with the strongest association. We validate its effect using a quantitative complementation test with CRISPR/Cas9-generated null mutants in nonstandard wild accessions. In contrast to pleiotropic null mutants, only pollen numbers are significantly affected by natural allelic variants. These data support theoretical predictions that reduced investment in male gametes is advantageous in predominantly selfing species.

A Similar Genetic Architecture Underlies the Convergent Evolution of the Selfing Syndrome in Capsella.

Whether, and to what extent, phenotypic evolution follows predictable genetic paths remains an important question in evolutionary biology. Convergent evolution of similar characters provides a unique opportunity to address this question. The transition to selfing and the associated changes in flower morphology are among the most prominent examples of repeated evolution in plants. In this study, we take advantage of the independent transitions to self-fertilization in the genus Capsella to compare the similarities between parallel modifications of floral traits and test for genetic and developmental constraints imposed on flower evolution in the context of the selfing syndrome. Capsella rubella and Capsella orientalis emerged independently but evolved almost identical flower characters. Not only is the evolutionary outcome identical but the same developmental strategies underlie the convergent reduction of flower size. This has been associated with convergent evolution of gene expression changes. The transcriptomic changes common to both selfing lineages are enriched in genes with low network connectivity and with organ-specific expression patterns. Comparative genetic mapping also suggests that, at least in the case of petal size evolution, these similarities have a similar genetic basis. Based on these results, we hypothesize that the limited availability of low-pleiotropy paths predetermines closely related species to similar evolutionary outcomes.

Standing genetic variation in a tissue-specific enhancer underlies selfing-syndrome evolution in Capsella.

Mating system shifts recurrently drive specific changes in organ dimensions. The shift in mating system from out-breeding to selfing is one of the most frequent evolutionary transitions in flowering plants and is often associated with an organ-specific reduction in flower size. However, the evolutionary paths along which polygenic traits, such as size, evolve are poorly understood. In particular, it is unclear how natural selection can specifically modulate the size of one organ despite the pleiotropic action of most known growth regulators. Here, we demonstrate that allelic variation in the intron of a general growth regulator contributed to the specific reduction of petal size after the transition to selfing in the genus Capsella Variation within this intron affects an organ-specific enhancer that regulates the level of STERILE APETALA (SAP) protein in the developing petals. The resulting decrease in SAP activity leads to a shortening of the cell proliferation period and reduced number of petal cells. The absence of private polymorphisms at the causal region in the selfing species suggests that the small-petal allele was captured from standing genetic variation in the ancestral out-crossing population. Petal-size variation in the current out-crossing population indicates that several small-effect mutations have contributed to reduce petal-size. These data demonstrate how tissue-specific regulatory elements in pleiotropic genes contribute to organ-specific evolution. In addition, they provide a plausible evolutionary explanation for the rapid evolution of flower size after the out-breeding-to-selfing transition based on additive effects of segregating alleles.

Divergent sorting of a balanced ancestral polymorphism underlies the establishment of gene-flow barriers in Capsella.

In the Bateson-Dobzhansky-Muller model of genetic incompatibilities post-zygotic gene-flow barriers arise by fixation of novel alleles at interacting loci in separated populations. Many such incompatibilities are polymorphic in plants, implying an important role for genetic drift or balancing selection in their origin and evolution. Here we show that NPR1 and RPP5 loci cause a genetic incompatibility between the incipient species Capsella grandiflora and C. rubella, and the more distantly related C. rubella and C. orientalis. The incompatible RPP5 allele results from a mutation in C. rubella, while the incompatible NPR1 allele is frequent in the ancestral C. grandiflora. Compatible and incompatible NPR1 haplotypes are maintained by balancing selection in C. grandiflora, and were divergently sorted into the derived C. rubella and C. orientalis. Thus, by maintaining differentiated alleles at high frequencies, balancing selection on ancestral polymorphisms can facilitate establishing gene-flow barriers between derived populations through lineage sorting of the alternative alleles.

Repeated evolutionary changes of leaf morphology caused by mutations to a homeobox gene.

Elucidating the genetic basis of morphological changes in evolution remains a major challenge in biology. Repeated independent trait changes are of particular interest because they can indicate adaptation in different lineages or genetic and developmental constraints on generating morphological variation. In animals, changes to "hot spot" genes with minimal pleiotropy and large phenotypic effects underlie many cases of repeated morphological transitions. By contrast, only few such genes have been identified from plants, limiting cross-kingdom comparisons of the principles of morphological evolution. Here, we demonstrate that the REDUCED COMPLEXITY (RCO) locus underlies more than one naturally evolved change in leaf shape in the Brassicaceae. We show that the difference in leaf margin dissection between the sister species Capsella rubella and Capsella grandiflora is caused by cis-regulatory variation in the homeobox gene RCO-A, which alters its activity in the developing lobes of the leaf. Population genetic analyses in the ancestral C. grandiflora indicate that the more-active C. rubella haplotype is derived from a now rare or lost C. grandiflora haplotype via additional mutations. In Arabidopsis thaliana, the deletion of the RCO-A and RCO-B genes has contributed to its evolutionarily derived smooth leaf margin, suggesting the RCO locus as a candidate for an evolutionary hot spot. We also find that temperature-responsive expression of RCO-A can explain the phenotypic plasticity of leaf shape to ambient temperature in Capsella, suggesting a molecular basis for the well-known negative correlation between temperature and leaf margin dissection.