Floral scent is important to plant reproduction because it attracts pollinators to the sexual organs. Therefore, volatile emission is usually tuned to the foraging activity of the pollinators. In Petunia hybrida, volatile benzenoids determine the floral aroma. Although the pathways for benzenoid biosynthesis have been characterized, the enzymes involved are less well understood. How production and emission are regulated is unknown. By targeted transcriptome analyses, we identified ODORANT1 (ODO1), a member of the R2R3-type MYB family, as a candidate for the regulation of volatile benzenoids in Petunia hybrida cv W115 (Mitchell) flowers. These flowers are only fragrant in the evening and at night. Transcript levels of ODO1 increased before the onset of volatile emission and decreased when volatile emission declined. Downregulation of ODO1 in transgenic P. hybrida Mitchell plants strongly reduced volatile benzenoid levels through decreased synthesis of precursors from the shikimate pathway. The transcript levels of several genes in this pathway were reduced by suppression of ODO1 expression. Moreover, ODO1 could activate the promoter of the 5-enol-pyruvylshikimate-3-phosphate synthase gene. Flower pigmentation, which is furnished from the same shikimate precursors, was not influenced because color and scent biosynthesis occur at different developmental stages. Our studies identify ODO1 as a key regulator of floral scent biosynthesis.

Anthocyanins and volatile phenylpropenes (isoeugenol and eugenol) in petunia (Petunia hybrida) flowers have the precursor 4-coumaryl coenzyme A (CoA) in common. These phenolics are produced at different stages during flower development. Anthocyanins are synthesized during early stages of flower development and sequestered in vacuoles during the lifespan of the flowers. The production of isoeugenol and eugenol starts when flowers open and peaks after anthesis. To elucidate additional biochemical steps toward (iso)eugenol production, we cloned and characterized a caffeoyl-coenzyme A O-methyltransferase (PhCCoAOMT1) from the petals of the fragrant petunia ‘Mitchell’. Recombinant PhCCoAOMT1 indeed catalyzed the methylation of caffeoyl-CoA to produce feruloyl CoA. Silencing of PhCCoAOMT1 resulted in a reduction of eugenol production but not of isoeugenol. Unexpectedly, the transgenic plants had purple-colored leaves and pink flowers, despite the fact that cv Mitchell lacks the functional R2R3-MYB master regulator ANTHOCYANIN2 and has normally white flowers. Our results indicate that down-regulation of PhCCoAOMT1 activated the anthocyanin pathway through the R2R3-MYBs PURPLE HAZE (PHZ) and DEEP PURPLE, with predominantly petunidin accumulating. Feeding cv Mitchell flowers with caffeic acid induced PHZ expression, suggesting that the metabolic perturbation of the phenylpropanoid pathway underlies the activation of the anthocyanin pathway. Our results demonstrate a role for PhCCoAOMT1 in phenylpropene production and reveal a link between PhCCoAOMT1 and anthocyanin production.