Regulation of class III peroxidases and respiratory burst oxidase homologs by biotic and abiotic stress in maize (Zea mays L.)

Abstract

The plasma membrane-bound peroxidases (PRX) zmprx01, zmprx66 and zmprx70 and the respiratory burst oxidase homologs (RBOH) rbohA, rbohB, rbohC and rbohD were analysed in this study. The distribution of the genes inside the roots was investigated by real-time qPCR. Therefor four different segments (root tip, elongation zone, differentiation zone and lateral roots) were in focus of the analyses. It could be observed that the genes are differently distributed in the root. The peroxidases were predominantly expressed in the elongation zone and almost not in the root tip. The rboh genes were more inhomogeneous distributed. For each RBOH a specific expression pattern could be detected. rbohA was mostly expressed in the differentiation zone. rbohB was more even expressed in the root. rbohC was even distributed as well but predominantly in the elongation zone. rbohD was mostly expressed in the differentiation zone. For a further investigation of the peroxidases plants were exposed to cadmium (short term and long term trial). The plants grew in cadmium contaminated hydroponics. zmprx66 and zmprx70 were upregulated after 15 minutes (quick response). Subsequently the expression went back to normal. Through the long term trial a decrease of each peroxidase was detected after three days of exposure. RNAi mutants were produced to analyse the lack of each peroxidase. RNAi was mediated by a heat shock inducible RNAi construct with double opposing promoters. This experiment was not finished, yet. By now it could be concluded that the down-regulation of zmprx66 decelerated the development of the whole plant. Further investigations are necessary. To find out more about the triggers for each gene and correlations between protein and mRNA abundance a stress profiling experiment was performed in accordance to the proteomic approach of Mika et al., 2010. The plants grew in hydroponics while the stress factors (chitosan, H2O2, NaCl, salicylic acid) were applied into the nutrition media. Additionally, mechanical wounding was performed. By the stress profiling it could be concluded that every gene has different triggers. The expression of the peroxidases did decrease by almost every treatment except zmprx70, which was positively affected by salicylic acid and wounding. Results suggest no correlation between protein abundance and mRNA, after 1 h or more. rbohC and rbohD were upregulated by H2O2, NaCl affected rbohB, rbohC and rbohD positively. Salicylic acid almost did not affect any RBOH except rbohC, which was slightly upregulated. rbohD was significantly upregulated through wounding. In association with Dr Meisrimler the genes were analysed under the impact of waterlogging. Waterlogging was performed with 28 days old potted maize plants. Mature and immature leaves were analysed separately. At that developmental stage it could be detected, that zmprx66 and zmprx70 were not expressed in leaves (control individual were analysed, preliminarily) but the protein (ZmPrx66) was found in that tissue. In this case no qPCR studies for zmprx66 and zmprx70 could be performed. It was observed that zmprx01 was predominantly expressed in immature leaves. The waterlogging had an impact on mature leaves. The expression was increased. For rbohB the same observation was made. The remaining RBOH seemed not to be affected by waterlogging, significantly. Because of this study many new information could be gained for zmprx01, zmprx66 and zmprx70 and rbohA, rbohB, rbohC and rbohD. Their triggers, co regulations and involvements in different processes could be identified or more clarified.

Die Plasmamembran gebundenen Peroxidasen zmprx01 zmprx66 und zmprx70 und die respiratory burst oxidase homologe (RBOH) rbohA, rbohB, rbohC, rbohD wurden in dieser Studie untersucht. Die Verteilung der Gene in den Wurzlen wurde über rela time PCR analysiert. Es wurden die Segmente Wurzelspitze, Elongationszone, Differenzierungszone und Lateralwurzeln untersucht. Weiter wurde die Expression der Gene durch biotischem und abiotischem Stress untersucht. Es wurde mit den Stressfaktoren Cadmium, Überflutung, Salz, H2O2, Chitosan experimentiert. Zur weiteren Analyse wurden die Peroxidasen durch RNAi herunterreguliert.