Rerouting the plant phenylpropanoid pathway by expression of a novel bacterial enoyl-CoA hydratase/lyase enzyme function

The gene for a bacterial enoyl-CoA hydratase (crotonase) homolog (HCHL) pre viously shown to convert 4-coumaroyl-CoA, caffeoyl-CoA, and feruloyl-CoA to the corresponding hydroxybenzaldehydes in vitro provided an opportunity to subvert the plant phenylpropanoid pathway and channel carbon flux through 4-hydroxybenzaldehyde and the important flavor compound 4-hydroxy-3-methoxy benzaldehyde (vanillin), Expression of the Pseudomonas fluorescens AN103 HC HL gene in two generations of tobacco plants caused the development of phen otypic abnormalities, including stunting, interveinal chlorosis and senesce nce, curled leaf margins, low pollen production, and male sterility. In sec ond generation progeny, the phenotype segregated with the transgene and tra nsgenic siblings exhibited orange/red coloration of the vascular ring, dist orted cells in the xylem and phloem bundles, and lignin modification/reduct ion. There was depletion of the principal phenolics concomitant with massiv e accumulation of novel metabolites, including the glucosides and glucose e sters of 4-hydroxybenzoic acid and vanillic acid and the glucosides of 4-hy droxybenzyl alcohol and vanillyl alcohol, HCHL plants exhibited increased a ccumulation of transcripts for phenylalanine ammonia-lyase, cinnamate-4-hyd roxylase, and 4-coumarate:CoA ligase, whereas beta -1,3-glucanase was suppr essed, This study, exploiting the ability of a bacterial gene to divert pla nt secondary metabolism, provides insight into how plants modify inappropri ately accumulated metabolites and reveals the consequences of depleting the major phenolic pools.