Supergene evolution via stepwise duplications and neofunctionalization of a floral-organ identity gene.

Heterostyly represents a fascinating adaptation to promote outbreeding in plants that evolved multiple times independently. While l-morph individuals form flowers with long styles, short anthers, and small pollen grains, S-morph individuals have flowers with short styles, long anthers, and large pollen grains. The difference between the morphs is controlled by an -locus "supergene" consisting of several distinct genes that determine different traits of the syndrome and are held together, because recombination between them is suppressed. In , the locus is a roughly 300-kb hemizygous region containing five predicted genes. However, with one exception, their roles remain unclear, as does the evolutionary buildup of the locus. Here we demonstrate that the MADS-box () gene at the locus determines anther position. In S-morph plants, promotes growth by cell expansion in the fused tube of petals and stamen filaments beneath the anther insertion point; by contrast, neither pollen size nor male incompatibility is affected by activity. The paralogue , from which arose by duplication, has maintained the ancestral B-class function in specifying petal and stamen identity, indicating that underwent neofunctionalization, likely at the level of the encoded protein. Genetic mapping and phylogenetic analysis indicate that the duplications giving rise to the style-length-determining gene and to occurred sequentially, with the duplication likely the first. Together these results provide the most detailed insight into the assembly of a plant supergene yet and have important implications for the evolution of heterostyly.