Glucose conjugation of anthranilate by the Arabidopsis UGT74F2 glucosyltransferase is required for tryptophan mutant blue fluorescence

Abstract

Plant mutants with defects in intermediate enzymes of the tryptophan biosynthetic pathway often display a blue fluorescent phenotype. This phenotype results from the accumulation of the fluorescent tryptophan precursor anthranilate, the bulk of which is found in a glucose-conjugated form. To elucidate factors that control fluorescent tryptophan metabolites, we conducted a genetic screen for suppressors of blue fluorescence in the Arabidopsis trp1-100 mutant, which has a defect in the second enzymatic step of the tryptophan pathway. This screen yielded loss-of-function mutations in the UDP-glucosyltransferase gene UGT74F2. The bacterially expressed UGT74F2 enzyme catalyzed a conjugation reaction, with free anthranilate and UDP-glucose as substrates, that yielded the same fluorescent glucose ester compound as extracted from the trp1-100 mutant. These results indicate that sugar conjugation of anthranilate by UGT74F2 allows its stable accumulation in plant tissues. A highly related Arabidopsis enzyme UGT74F1 could also catalyze this reaction in vitro and could complement the ugt74F2 mutation when overexpressed in vivo. However, the UGT74F1 gene is expressed at a lower level than the UGT74F2 gene. Therefore, even though UGT74F1 and UGT74F2 have redundant conjugating activities toward anthranilate, UGT74F2 is the major source of this activity in the plant.