| The tryptophan (trp) pathway in Arabidopsis thaliana provides the amino acid trp for protein synthesis and important secondary metabolites such as the plant hormone indole-3-acetic acid and defense compounds camalexin and indole glucosinolates. These metabolites participate in the regulation of growth and development, and protection against pathogenic bacteria and predatory insects. The trp pathway is subject to strict regulatory mechanisms that ensure proper spatial and temporal production of trp and trp-derived compounds.; This thesis describes two different approaches to understanding regulation of the trp pathway. First, I conducted a genetic screen to elucidate the homeostasis of the first intermediate compound in the trp pathway, anthranilate. In the trp1-100 mutant strain, anthranilate accumulates in a glucose-conjugated form conferring a blue fluorescent phenotype. I isolated mutations in the Arabidopsis UGT74F2 glucosyltransferase gene as suppressors of trp1-100 blue fluorescence, and showed that the heterologously-expressed UGT74F2 enzyme catalyzes a conjugation reaction using UDP-glucose and anthranilate as substrates to yield a glucose ester in vitro. Thus, glucose conjugation by UGT74F2 is necessary for the stable accumulation of anthranilate in the plant.; Previous work identified an overexpression allele of the ATR1 Myb transcription factor gene as a trp pathway activating mutation. To further characterize ATR1, I monitored its expression patterns in the plant, and assayed the consequences of high constitutive ATR1 overexpression via a transgene construct. I also isolated and characterized a loss-of-function mutation in the ATR1 gene. These experiments show that ATR1 is necessary for upregulation of trp pathway genes, particularly genes encoding secondary metabolism enzymes for indole glucosinolate biosynthesis. Because indole glucosinolates are key anti-insect compounds in plants of the mustard family, modulation of ATR1 levels in agricultural plants of this family has the potential to yield crops with improved insect resistance. |
