ABSTRACT
Floral volatile benzenoid/phenylpropanoid (FVBP) biosynthesis is a complex and coordinate cellular process executed by petal limb cells of a Petunia×hybrida cv. 'Mitchell Diploid' (MD) plant. In MD flowers, the majority of benzenoid volatile compounds are derived from a core phenylpropanoid pathway intermediate by a coenzyme A (CoA) dependent, ß-oxidative scheme. Metabolic flux analysis, reverse genetics, and biochemical characterizations of key enzymes in this pathway have supported this putative concept. However, the theoretical first enzymatic reaction, which leads to the production of cinnamoyl-CoA, has only been physically demonstrated in a select number of bacteria like Streptomyces maritimus through mutagenesis and recombinant protein production. A transcript has been cloned and characterized from MD flowers that shares high homology with an Arabidopsis thaliana transcript ACYL-ACTIVATING ENZYME11 (AtAAE11) and the S. maritimus ACYL-COA:LIGASE (SmEncH). In MD, the PhAAE transcript accumulates in a very similar manner as bona fide FVBP network genes, i.e. high levels in an open flower petal and ethylene regulated. In planta, PhAAE is localized to the peroxisome. Upon reduction of PhAAE transcript through a stable RNAi approach, transgenic flowers emitted a reduced level of all benzenoid volatile compounds. Together, the data suggest that PhAAE may be responsible for the activation of t-cinnamic acid, which would be required for floral volatile benzenoid production in MD.
Subject(s)
Benzene Derivatives/metabolism , Flowers/enzymology , Peroxisomes/enzymology , Petunia/enzymology , Plant Proteins/metabolism , Propanols/metabolism , Amino Acid Sequence , DNA, Plant/chemistry , DNA, Plant/genetics , Flowers/chemistry , Flowers/genetics , Flowers/ultrastructure , Gene Expression Regulation, Plant , Metabolic Networks and Pathways , Microscopy, Confocal , Molecular Sequence Data , Petunia/chemistry , Petunia/genetics , Petunia/ultrastructure , Phylogeny , Plant Proteins/genetics , Plant Roots/chemistry , Plant Roots/enzymology , Plant Roots/genetics , Plant Roots/ultrastructure , Plant Stems/chemistry , Plant Stems/enzymology , Plant Stems/genetics , Plant Stems/ultrastructure , RNA Interference , RNA, Messenger/genetics , RNA, Plant/genetics , Recombinant Proteins , Sequence AlignmentABSTRACT
R2R3-MYB transcription factors (TFs) are involved in diverse aspects of plant biology. Recently an R2R3-MYB was identified in Petunia x hybrida line P720 to have a role in the transcriptional regulation of floral volatile production. We propose a more foundational role for the R2R3-MYB TF EMISSION OF BENZENOIDS II (EOBII). The homolog of EOBII was isolated and characterized from P. x hybrida 'Mitchell Diploid' (MD) and Nicotiana attenuata. For both MD and N. attenuata, EOBII transcript accumulates to high levels in floral tissue with maximum accumulation at flower opening. When EOBII transcript levels are severely reduced using a stable RNAi (ir) approach in MD and N. attenuata, ir-EOBII flowers fail to enter anthesis and prematurely senesce. Transcript accumulation analysis demonstrated core phenylpropanoid pathway transcripts and cell wall modifier transcript levels are altered in ir-EOBII flowers. These flowers can be partially complemented by feeding with a sucrose, t-cinnamic acid, and gibberellic acid solution; presumably restoring cellular aspects sufficient for flower opening. Additionally, if ethylene sensitivity is blocked in either MD or N. attenuata, ir-EOBII flowers enter anthesis. These experiments demonstrate one R2R3-MYB TF can control a highly dynamic process fundamental to sexual reproduction in angiosperms: the opening of flowers.
