The genetic basis of replicated bullseye pattern reduction across the Trionum Complex

Angiosperm flowers exhibit a wide diversity of colorful motifs on their petals. Such patterns fulfill both biotic and abiotic functions, mediating plant-pollinator communication and providing protection against damaging UV rays or desiccation. These motifs are often evolutionary labile, varying in size, shape and hue between closely related species and constitute excellent systems to illuminate the evolutionary processes that generate morphological diversity or instead lead to the repetitive emergence of similar forms. Hibiscus trionum flowers have a prominent bullseye pattern combining a purple center contrasting against a white margin. H. trionum belongs to a small clade of Hibiscus known as the Trionum Complex that displays a range of petal patterns within and between species. Here, we integrate phylogenomic approaches, molecular techniques and genetic manipulation to solve species relationships across the Trionum Complex, identify key genes involved in the production of bullseye pigmentation, and reveal molecular events underpinning pattern variation during the evolution of the group. We find that repetitive bullseye reduction events primarily occur through independent modifications of a single genetic locus encoding BERRY1, an R2R3 MYB that regulates anthocyanin pigment production in petals. Moreover, we demonstrate that buff-tailed bumblebees (Bombus terrestris) discriminate against flowers with smaller bullseye sizes suggesting that a reduction in bullseye proportions potentially impacts plant fitness and contributes to trait distribution across the group. Collectively, our results demonstrate how repeated mutations in a single locus led to morphological variation in petal patterning, a trait that contributes to angiosperm reproductive isolation and speciation.