NSF Award
0095075
The end products of the phenylpropanoid pathway play important roles in plant structure and plant growth and development, as well as in plants' defenses against biotic and abiotic stresses. Phenylpropanoid pathway-derived metabolites are important in both human health and in agricultural contexts. An important gap in our understanding of phenylpropanoid metabolism is represented by p-coumarate 3-hydroxylase (C3H), the only enzyme of the pathway that has not been characterized, and the enzyme encoded by the only gene of the pathway that has not been cloned. The 3-hydroxylase is an enigmatic enzyme. It has eluded all attempts over the last thirty years to unambiguously characterize it in detail at the enzymatic level. This research project focuses on a new mutant of Arabidopsis that is likely to be defective in the gene encoding this enzyme.<br/>Past research has shown that Arabidopsis is an excellent model plant for the isolation of previously unidentified phenylpropanoid pathway genes. By screening for plants that fail to accumulate soluble fluorescent phenylpropanoid secondary metabolites, a number of new Arabidopsis mutants that display a reduced epidermal fluorescence (ref) phenotype have been identified. One of these mutants, ref8, is defective in C3H activity. The identification of the ref8 mutant provides a unique opportunity to isolate the gene encoding C3H. A preliminary phenotypic characterization of the ref8 mutant has been conducted, and the C3H gene has been cloned. C3H is a cytochrome P450-dependent monooxygenase, the activity of which has been verified by expression of the wild-type gene in yeast. The goals of this project are to characterize this poorly-studied enzyme, to clarify its role in the phenylpropanoid pathway, to identify when and where the protein is expressed, and to determine why ref8 mutations lead to alterations in plant growth and development.
