Abstract
Class III plant peroxidases (Prxs) are involved in the oxidative polymerization of lignins. Zinnia elegans Jacq. Basic peroxidase (ZePrx) has been previously characterized as capable of catalyzing this reaction in vitro and the role in lignin biosynthesis of several of its Arabidopsis thaliana homologous has been previously confirmed. In the present work, ZePrx was overexpressed in Nicotiana tabacum to further characterize its function in planta with particular attention to its involvement in lignin biosynthesis. Since Prxs are known to alter ROS levels by using them as electron acceptor or producing them in their catalytic activity, the impact of this overexpression in redox homeostasis was studied by analyzing the metabolites and enzymes of the ascorbate-glutathione cycle. In relation to the modification induced by ZePrx overexpression in lignin composition and cellular metabolism, the carbohydrate composition of the cell wall as well as overall gene expression through RNA-Seq were analyzed. The obtained results indicate that the overexpression of ZePrx caused an increase in syringyl lignin in cell wall stems, suggesting that ZePrx is relevant for the oxidation of sinapyl alcohol during lignin biosynthesis, coherently with its S-peroxidase nature. The increase in the glucose content of the cell wall and the reduction of the expression of several genes involved in secondary cell wall biosynthesis suggests the occurrence of a possible compensatory response to maintain cell wall properties. The perturbation of cellular redox homeostasis occurring as a consequence of ZePrx overexpression was kept under control by an increase in APX activity and a reduction in ascorbate redox state. In conclusion, our results confirm the role of ZePrx in lignin biosynthesis and highlight that its activity alters cellular pathways putatively aimed at maintaining redox homeostasis.
Highlights
Peroxidases (EC 1.11.1) are enzymes capable of oxidize many substrates using hydrogen peroxide (H2O2) as co-substrate (De Gara, 2004; Oliva et al, 2009)
Overexpressing Zinnia elegans Jacq. Basic peroxidase (ZePrx) Transformed Lines Accumulated High Level of an Active Protein N. tabacum plants were transformed with the ZePrx coding sequence under the control of the strong and constitutive CaMV35S promoter and eleven stable independent transformed lines were obtained
Since ZePrx has been reported to catalyze the last step of lignin biosynthesis (Gabaldón et al, 2005), the impact of its overexpression was firstly investigated in the stem, an organ in which the lignification is pivotal, especially during its secondary growth
Summary
Peroxidases (EC 1.11.1) are enzymes capable of oxidize many substrates using hydrogen peroxide (H2O2) as co-substrate (De Gara, 2004; Oliva et al, 2009). Prxs are apoplastic and vacuolar plant peroxidases presenting a double catalytic-cycle, which allow them to both consume and produce reactive oxygen species (ROS; Chen and Schopfer, 1999; Zámocky and Obinger, 2010). Several reverse genetic studies, focused on the catalytic properties of these enzymes as well as on their specialized function, demonstrate that Prxs can be involved in a diverse range of functions (reviewed in Shigeto and Tsutstumi, 2016). The diverse functions of Prxs are accomplished through the oxidation of specific molecules such as lignin precursors during lignin polymerization (Sasaki et al, 2004; Gabaldón et al, 2006) and/or through the regulation of specific ROS levels (Kawaoka et al, 2003; Liszkay et al, 2003; Kim et al, 2008; Sundaravelpandian et al, 2013; Peters et al, 2017). One of the aspects drawing more attention is Prx involvement in lignification (reviewed in Marjamaa et al, 2009)
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