PhDAHP1 is required for floral volatile benzenoid/phenylpropanoid biosynthesis in Petunia × hybrida cv ‘Mitchell Diploid’

Abstract

Floral volatile benzenoid/phenylpropanoid (FVBP) biosynthesis consists of numerous enzymatic and regulatory processes. The initial enzymatic step bridging primary metabolism to secondary metabolism is the condensation of phosphoenolpyruvate (PEP) and erythrose-4-phosphate (E4P) carried out via 3-DEOXY-D-ARABINO-HEPTULOSONATE-7-PHOSPHATE (DAHP) synthase. Here, identified, cloned, localized, and functionally characterized were two DAHP synthases from the model plant species Petunia × hybrida cv ‘Mitchell Diploid’ (MD). Full-length transcript sequences for PhDAHP1 and PhDAHP2 were identified and cloned using cDNA SMART libraries constructed from pooled MD corolla and leaf total RNA. Predicted amino acid sequence of PhDAHP1 and PhDAHP2 proteins were 76% and 80% identical to AtDAHP1 and AtDAHP2 from Arabidopsis, respectively. PhDAHP1 transcript accumulated to relatively highest levels in petal limb and tube tissues, while PhDAHP2 accumulated to highest levels in leaf and stem tissues. Through floral development, PhDAHP1 transcript accumulated to highest levels during open flower stages, and PhDAHP2 transcript remained constitutive throughout. Radiolabeled PhDAHP1 and PhDAHP2 proteins localized to plastids, however, PhDAHP2 localization appeared less efficient. PhDAHP1 RNAi knockdown petunia lines were reduced in total FVBP emission compared to MD, while PhDAHP2 RNAi lines emitted ‘wildtype’ FVBP levels. These results demonstrate that PhDAHP1 is the principal DAHP synthase protein responsible for the coupling of metabolites from primary metabolism to secondary metabolism, and the ultimate biosynthesis of FVBPs in the MD flower.