Summary: Within the angiosperm order Caryophyllales, an unusual class of pigments known as betalains can replace the otherwise ubiquitous anthocyanins. In contrast to the phenylalanine‐derived anthocyanins, betalains are tyrosine‐derived pigments which contain the chromophore betalamic acid. The origin of betalain pigments within Caryophyllales and their mutual exclusion with anthocyanin pigments have been the subject of considerable research. In recent years, numerous discoveries, accelerated by ‐omic scale data, phylogenetics and synthetic biology, have shed light on the evolution of the betalain biosynthetic pathway in Caryophyllales. These advances include the elucidation of the biosynthetic steps in the betalain pathway, identification of transcriptional regulators of betalain synthesis, resolution of the phylogenetic history of key genes, and insight into a role for modulation of primary metabolism in betalain synthesis. Here we review how molecular genetics have advanced our understanding of the betalain biosynthetic pathway, and discuss the impact of phylogenetics in revealing its evolutionary history. In light of these insights, we explore our new understanding of the origin of betalains, the mutual exclusion of betalains and anthocyanins, and the homoplastic distribution of betalain pigmentation within Caryophyllales. We conclude with a speculative conceptual model for the stepwise emergence of betalain pigmentation.

A major function of angiosperm flowers is the recruitment of animal pollinators that serve to transfer pollen among conspecific plants. Distinct sets of floral characteristics, called pollination syndromes, are correlated with visitation by specific groups of pollinators. Switches among pollination syndromes have occurred in many plant families. Such switches must have involved coordinated changes in multiple traits and multiple genes. Two well-studied floral traits affecting pollinator attraction are petal color and scent production. We review current knowledge about the biosynthetic pathways for floral color and scent production and their interaction at the genetic and biochemical levels. A key question in the field concerns the genes that underlie natural variation in color and scent and how such genes affect pollinator preference, reproductive isolation, and ultimately speciation.

Summary: The evolution of l‐DOPA 4,5‐dioxygenase activity, encoded by the gene DODA, was a key step in the origin of betalain biosynthesis in Caryophyllales. We previously proposed that l‐DOPA 4,5‐dioxygenase activity evolved via a single Caryophyllales‐specific neofunctionalisation event within the DODA gene lineage. However, this neofunctionalisation event has not been confirmed and the DODA gene lineage exhibits numerous gene duplication events, whose evolutionary significance is unclear. To address this, we functionally characterised 23 distinct DODA proteins for l‐DOPA 4,5‐dioxygenase activity, from four betalain‐pigmented and five anthocyanin‐pigmented species, representing key evolutionary transitions across Caryophyllales. By mapping these functional data to an updated DODA phylogeny, we then explored the evolution of l‐DOPA 4,5‐dioxygenase activity. We find that low l‐DOPA 4,5‐dioxygenase activity is distributed across the DODA gene lineage. In this context, repeated gene duplication events within the DODA gene lineage give rise to polyphyletic occurrences of elevated l‐DOPA 4,5‐dioxygenase activity, accompanied by convergent shifts in key functional residues and distinct genomic patterns of micro‐synteny. In the context of an updated organismal phylogeny and newly inferred pigment reconstructions, we argue that repeated convergent acquisition of elevated l‐DOPA 4,5‐dioxygenase activity is consistent with recurrent specialisation to betalain synthesis in Caryophyllales.

Analysis of mutants with increased branching has revealed the strigolactone synthesis/perception pathway which regulates branching in plants. However, whether variation in this well conserved developmental signaling system contributes to the unique plant architectures of different species is yet to be determined. We examined petunia orthologs of the Arabidopsis MAX1 and MAX2 genes to characterize their role in petunia architecture. A single ortholog of MAX1, PhMAX1 which encodes a cytochrome P450, was identified and was able to complement the max1 mutant of Arabidopsis. Petunia has two copies of the MAX2 gene, PhMAX2A and PhMAX2B which encode F-Box proteins. Differences in the transcript levels of these two MAX2-like genes suggest diverging functions. Unlike PhMAX2B, PhMAX2A mRNA levels change in leaves of differing age/position on the plant. Nonetheless, this gene functionally complements the Arabidopsis max2 mutant indicating that the biochemical activity of the PhMAX2A protein is not significantly different from MAX2. The expression of the petunia strigolactone pathway genes (PhCCD7, PhCCD8, PhMAX1, PhMAX2A, and PhMAX2B) was then further investigated throughout the development of wild-type petunia plants. Three of these genes showed changes in mRNA levels over a development series. Alterations to the expression patterns of these genes may influence the branching growth habit of plants by changing strigolactone production and/or sensitivity. These changes could allow both subtle and dramatic changes to branching within and between species.