Analysis of the expression and function of polyphenol oxidase in hybrid poplar

In order to probe the physiological functions and expression of polyphenol oxidase (PPO) in hybrid poplar (Populus trichocarpa x P. deltoides), full-length cDNAs of three tissue-specific PtdPPOs were obtained and characterized. Based on the results of northern blots using gene-specific probes, PtdPPO1 is a wound-inducible leaf PPO; PtdPPO2 is a constitutive and wounding-inducible PPO mainly found in conducting organs; and PtdPPO3 is root-specific and does not respond to stresses tested. Distinct distribution among hybrid poplar organs in addition to their differential responses to wounding and defense elicitors indicate that these three PPOs may perform different physiological roles in hybrid poplar. In order to directly test the defensive role of PPO in herbivore defense, PtdPPO1-overexpressing hybrid aspen (Populus tremula x P. alba) was constructed and forest tent caterpillar (FTC) was tested for their survivorship and weight gain when reared on those transgenics. We found that PPO1 negatively affected the growth and survivorship of FTC when the FTC larvae were hatched from old egg mass, which had low vigor. This confirmed the anti-herbivore role of PPO1 in hybrid poplar. In order to investigate the biochemical properties of hybrid poplar PPOs, PPO1 and PPO2 proteins were partially purified through hydrophobic chromatography. Biochemical analysis showed that these two PPO proteins exhibited somewhat different enzymatic properties such as storage stability, heat sensibility, SDS activation, pH optima and substrate preferences. In summary, this work thoroughly investigated the expression and induction patterns of different PPO genes in hybrid poplar, directly confirmed the defensive role of PPO1 in Populus herbivore defense and revealed that the three different PPO proteins in hybrid poplar possess distinct enzymatic properties.