Quantitative relationship between phenylalanine ammonia-lyase levels and phenylpropanoid accumulation in transgenic tobacco identifies a rate-determining step in natural product synthesis

Abstract

Phenylalanine ammonia-lyase (PAL) catalyzes the first step in phenylpropanoid synthesis. The role of PAL in pathway regulation was investigated by measurement of product accumulation as a function of enzyme activity in a collection of near-isogenic transgenic tobacco plants exhibiting a range of PAL levels from wild type to 0.2% of wild type. In leaf tissue, PAL level is the dominant factor regulating accumulation of the major product chlorogenic acid and overall flux into the pathway. In stems, PAL at wild-type levels contributes, together with downstream steps, in the regulation of lignin deposition and becomes the dominant, rate-determining step at levels 3- to 4-fold below wild type. The metabolic impact of elevated PAL levels was investigated in transgenic leaf callus that overexpressed PAL. Accumulation of the flavonoid rutin, the major product in wild-type callus, was not increased, but several other products accumulated to similarly high levels. These data indicate that PAL is a key step in the regulation of overall flux into the pathway and, hence, accumulation of major phenylpropanoid products, with the regulatory architecture of the pathway poised so that downstream steps control partitioning into different branch pathways.