Abstract
Camptothecin (CPT) has unique anti-tumor activities, and its analogs such as irinotecan and topotecan are clinically used as anti-cancer drugs. Recently, CPT biosynthesis pathway is gradually being resolved. However, few studies have been conducted on transcription factors (TFs), which regulate CPT biosynthesis in Ophiorrhiza pumila. In this study, we cloned and functionally identified a new OpWRKY1 TF from O. pumila that was preferentially expressed in stems and responded to exogenous elicitors such as gibberellin, acetylsalicylic acid, salicylic acid, abscisic acid and 1-aminocyclopropane-carboxylic acid. Subcellular localization assay showed that OpWRKY1 was specifically expressed in the nucleus. Transgenic results revealed that over-expression of OpWRKY1 significantly repressed CPT accumulation in the hairy roots by inhibiting the expression of several CPT biosynthesis genes, especially OpCPR. Furthermore, Dual-Luciferase assay and yeast-one-hybrid assay showed that OpCPR was a target of OpWRKY1. The above results elucidated how OpWRKY1 directly down-regulated the transcription of OpCPR and inhibited CPT biosynthesis in O. pumila. Our findings indicate that OpWRKY1 acts as a negative regulator of CPT biosynthesis in O. pumila and provide new insights on metabolic engineering of CPT metabolism in the future.
Key message
Overexpression of OpWRKY1 inhibits CPT accumulation in transgenic O. pumila hairy roots by reducing the expression of CPT biosynthetic gene OpCPR.
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Acknowledgements
This work was funded by National Natural Science Fund of China (31571735, 81522049, 31270007), Zhejiang Provincial Ten Thousands Program for Leading Talents of Science and Technology Innovation (2018R52050), Zhejiang Provincial Program for the Cultivation of High-level Innovative Health talents, Shanghai Science and Technology Committee Project (17JC1404300), and Opening project of Zhejiang provincial preponderant and characteristic subject of Key University (Traditional Chinese Pharmacology), Zhejiang Chinese Medical University (ZYAOX2018017). Besides, we thank LetPub (www.letpub.com) for its linguistic assistance during the preparation of this manuscript.
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GK designed the experiments; MX, CW (Chao Wu), LZ and CW (Can Wang) performed most of experiments; MX, CW (Chao Wu) and WZ analyzed the data; MX, YW, YM and GK wrote the manuscript.
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Xu, M., Wu, C., Zhao, L. et al. WRKY transcription factor OpWRKY1 acts as a negative regulator of camptothecin biosynthesis in Ophiorrhiza pumila hairy roots. Plant Cell Tiss Organ Cult 142, 69–78 (2020). https://doi.org/10.1007/s11240-020-01833-2
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DOI: https://doi.org/10.1007/s11240-020-01833-2