Herbicide Bioactivation in Arabidopsis Thaliana: Role of a Carboxylesterase

Arabidopsis thaliana contains multiple carboxyesterases (AtCXEs) with activities toward xenobiotics, including herbicide esters that are activated to their phytotoxic acids upon hydrolysis. On the basis of their susceptibility to inhibition by organophosphates, these AtCXEs are all serine hydrolases. Using a trifunctional probe (TriFP) bearing a fluorophosphonate together with biotin and rhodamine to facilitate detection and recovery, four dominant serine hydrolases were identified in the proteome of Arabidopsis. Using a combination of protein purification, capture with the TriFP and proteomics, one of these hydrolases, AtCXE12, was shown to be the major carboxylesterase responsible for hydrolysing the pro-herbicide methyl-2,4dichlorophenoxyacetate (2,4-D-methyl) to the phytotoxic acid 2,4-D. Recombinant expression of the other identified hydrolases showed that AtCXE12 was unique in hydrolyzing 2,4-D-methyl. To determine the importance of AtCXE12 in herbicide metabolism and efficacy, the respective TDNA knock-out (atcxe12) plants were characterized and shown to lack expression of AtCXE12 and have greatly reduced levels of 2,4-D-methyl hydrolyzing activity. Young atcxe12 seedlings were less sensitive than wild type plants to 2,4-D-methyl, confirming a role for the enzyme in herbicide bioactivation in Arabidopsis.