TheArabidopsis thalianaKnockout Mutant forPhytochelatin Synthase1(cad1-3) Is Defective in Callose Deposition, Bacterial Pathogen Defense and Auxin Content, But Shows an Increased Stem Lignification

The enzyme phytochelatin synthase (PCS) has long been studied with regard to its role in metal(loid) detoxification in several organisms, i.e., plants, yeasts, and nematodes. It is in fact widely recognized that PCS detoxifies a number of heavy metals by catalyzing the formation of thiol-rich oligomers, namely phytochelatins, from glutathione and related peptides. However, recent investigations have highlighted other possible roles played by the PCS enzyme in the plant cell, e.g., the control of pathogen-triggered callose deposition. In order to examine novel aspects ofArabidopsis thalianaPCS1 (AtPCS1) functions and to elucidate its possible roles in the secondary metabolism, metabolomic data ofA. thalianawild-type andcad1-3mutant were compared, the latter lacking AtPCS1. HPLC-ESI-MS analysis showed differences in the relative levels of metabolites from the glucosinolate and phenylpropanoid pathways betweencad1-3and wild-type plants. Specifically, in control (Cd-untreated) plants, higher levels of 4-methoxy-indol-3-ylmethylglucosinolate were found incad1-3plants vs. wild-type. Moreover, thecad1-3mutant showed to be impaired in the deposit of callose after Cd exposure, suggesting that AtPCS1 protects the plant against the toxicity of heavy metals not only by synthesizing PCs, but also by contributing to callose deposition. In line with the contribution of callose in counteracting Cd toxicity, we found that another callose-defective mutant,pen2-1, was more sensitive to high concentrations of Cd than wild-type plants. Moreover,cad1-3plants were more susceptible than wild-type to the hemibiotrophic bacterial pathogenPseudomonas syringae. The metabolome also revealed differences in the relative levels of hydroxycinnamic acids and flavonols, with consequences on cell wall properties and auxin content, respectively. First, increased lignification in thecad1-3stems was found, probably aimed at counteracting the entry of Cd into the inner tissues. Second, incad1-3shoots, increased relative levels of kaempferol 3,7 dirhamnoside and quercetin hexoside rhamnoside were detected. These flavonols are endogenous inhibitors of auxin transportin planta; auxin levels in both roots and shoots of thecad1-3mutant were in fact lower than those of the wild-type. Overall, our data highlight novel aspects of AtPCS1 functions inA. thaliana.