AUTHOR=Zhang Jie , Wu Zhaochen , Han Nuan , Wang Dongmei 

TITLE=Functional validation of ZbFAD2 and ZbFAD3 in the alkylamide biosynthesis pathway from Zanthoxylum bungeanum Maxim

JOURNAL=Frontiers in Plant Science

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2022.991882

DOI=10.3389/fpls.2022.991882

ISSN=1664-462X

ABSTRACT=<p>The spicy taste and medicinal properties of <italic>Zanthoxylum bungeanum</italic> are imparted by several alkylamides. Although most studies have focused on their isolation and identification, few have reported their biosynthesis pathways. Among the differentially expressed genes (DEGs) reported in the numerous varieties of <italic>Z. bungeanum</italic>, some might contribute to alkylamide biosynthesis. However, they are not yet functionally validated. The present study explored the function of two genes, <italic>ZbFAD2</italic> and <italic>ZbFAD3</italic>, in the alkylamide biosynthesis pathway, and their stable and transient expression in <italic>Arabidopsis thaliana</italic> and <italic>Nicotiana benthamiana</italic> were also analyzed. As compared with the wild-type (WT), the fatty acid content analysis indicated that <italic>ZbFAD2</italic>-<italic>A. thaliana</italic> transgenic seeds had lower oleic acid and higher linoleic acid contents, while the <italic>ZbFAD3</italic>-<italic>A. thaliana</italic> transgenic seeds showed lower linoleic acid and higher α-linolenic acid levels. Moreover, hydroxy-α-sanshool, a major alkylamide, was considerably higher in the <italic>ZbFAD2</italic>-<italic>N. benthamiana</italic> transgenic plants (0.2167 ± 0.0026 mg/g) than in the WT (0.0875 ± 0.0049 mg/g), while it was lower in the <italic>ZbFAD3</italic>-<italic>N. benthamiana</italic> transgenic plants (0.0535 ± 0.0037 mg/g). These results suggest that both <italic>ZbFAD2</italic> and <italic>ZbFAD3</italic> are vital alkylamide biosynthesis enzymes in <italic>Z. bungeanum</italic>. Our study not only helps to scale up the alkylamide production, but also establishes the role of the uncharacterized genes.</p>