Xylem Peroxidases: Purification and Altered Expression

ABSTRACT The peroxidase-dependent oxidation of the lignin monomer analogue syringaldazine (SYR) shows an appealing co-localization with lignification in most species analyzed. We have isolated two SYR-oxidizing peroxidases from the xylem of poplar ( P. trichocarpa ‘Trichobel’). We have shown that these were the only isoenzymes able to catalyze this reaction in the xylem, and that the corresponding activity correlates with actively lignifying cells within the poplar xylem. Furthermore, we have isolated a cDNA that codes for one of these enzymes (PXP 3-4) and demonstrated that the mRNA is expressed in the bark and the xylem of the stem and in the xylem of the roots. The cDNA encodes a peroxidase that is expressed as a preprotein with signalpeptides at both termini. The peroxidase cDNA PXP 3-4 was expressed in P. trenula × P. alba in sense orientation, using constructs encoding either the native enzyme or a version without the C-terminal propeptide. Also plants expressing this cDNA in antisense orientation were generated. Overexpressing lines with more than 800-fold higher peroxidase activity were identified. In the bark of these plants, half of the extracted proteins corresponded to PXP 3-4, which is more than 0.5 mg peroxidase per g of bark. All overexpressing lines were phenotypically normal. No alteration was observed in lignin amount, condensation or monolignol composition and the metabolic profiles and the redox state of these plants were unaltered.