1. Responses of defense signals, biosynthetic gene transcription and taxoid biosynthesis to elicitation by a novel synthetic jasmonate in cell cultures of Taxus chinensis
Fengxian Hu, Junhai Huang, Yufang Xu, Xuhong Qian, Jian-Jiang Zhong Biotechnol Bioeng. 2006 Aug 20;94(6):1064-71. doi: 10.1002/bit.20921.
Recently novel synthetic jasmonate derivatives were shown to have very powerful stimulating effects on the biosynthesis of taxuyunnanine C (Tc) by Taxus chinensis cells (Biotech Bioeng, 86:595; ibid. 86: 809, 2004). To provide an insight into the elicitation mechanism of the newly synthesized elicitors, by taking 2-hydroxyethyl jasmonate (HEJ, at 100 microM) as a typical example, in this work the defense signals were detected and their influences on the expression of important genes in taxoid biosynthetic pathway were examined in cell cultures of T. chinensis. The oxidative burst (induced H2O2 production) was confirmed, and the induction of lipoxygenase (LOX) activity and intracellular jasmonate acid (JA) synthesis was found. The gene transcription of geranylgeranyl diphosphate synthase (GGPPs) and taxa-4(5),11(12)-diene synthase (TS) was up-regulated by HEJ elicitation compared to control. Inhibition of JA biosynthesis by a putative LOX inhibitor, ibuprofen (IBU), effectively depressed the HEJ-induced up-regulation of GGPPs and TS gene transcription levels. In contrast, the inhibition of H2O2 production by membrane NADPH oxidase inhibitor, diphenylene iodonium (DPI), did not affect the transcriptions of those genes. For the HEJ-induced Tc production, it was suppressed with addition of DPI or IBU. The results suggest that both H2O2 and JA signals were involved in HEJ-induced Tc biosynthesis, and JA mediated the induction of GGPPs and TS genes expression, but H2O2 was not essential to activate them. Finally, a signal transduction cascade from defense signal response to activated transcription of taxoid biosynthetic genes and enhanced Tc production is proposed.
2. Quantitative influence of endogenous salicylic acid level on taxuyunnanine C biosynthesis in suspension cultures of Taxus chinensis
Xin Zhou, Jian-Jiang Zhong Biotechnol Bioeng. 2011 Jan;108(1):216-21. doi: 10.1002/bit.22915.
Intracellular signals are critical to secondary metabolite biosynthesis by plant cells, but their quantitative effects are not yet well understood in plant cell cultures. Using Taxus chinensis suspension culture, which has the potential to provide a sustainable supply of highly useful bioactive taxoids, this work investigated the impact of endogenous salicylic acid (SA) level on taxuyunnanine C (Tc) biosynthesis. It was observed that the Tc accumulation was strongly dependent on the endogenous SA level. Under elicitation with an endogenous SA of 97.1 ± 11.8 µg/g dry weight, a maximal Tc content of 10.3 ± 0.52 mg/g dry weight was obtained. The transcription levels of Tc biosynthetic genes of geranylgeranyl diphosphate synthase and taxadiene synthase were also analyzed, and they were increased with increase of internal SA level. The results demonstrated that manipulation of endogenous SA level could be an efficient strategy for improving secondary metabolite biosynthesis in plant cell culture.
3. Novel, unnatural benzo-1,2,3-thiadiazole-7-carboxylate elicitors of taxoid biosynthesis
Yufang Xu, Zhengjiang Zhao, Xuhong Qian, Zhigang Qian, Wenhong Tian, Jianjiang Zhong J Agric Food Chem. 2006 Nov 15;54(23):8793-8. doi: 10.1021/jf0618574.
In order to establish the chemical biological technology for production of valuable secondary metabolites, a novel family of unnatural elicitors derived from the plant activator benzo-1,2,3-thiadiazole-7-carboxylic acid were designed and synthesized. New synthetic elicitors that showed powerful eliciting activities upon taxoid biosynthesis by Taxus chinensis suspension cells were obtained. For example, benzo-1,2,3-thiadiazole-7-carboxylic acid 2-(2-hydroxybenzoxyl)ethyl ester was more effective and resulted in nearly 40% increase in taxuyunnanine C content and production in comparison with methyl jasmonate, which was previously reported as the most powerful chemical elicitor for taxoid biosynthesis. The novel class of elicitors was found to induce plant defense responses, including promotion of H(2)O(2) levels originating from oxidative burst and activation of phenylalanine ammonia lyase. Interestingly the plant defense responses induced corresponded well to the superior stimulating activity in T. chinensis cell cultures. The work indicates that the newly synthesized benzothiadiazoles can act as a new family of elicitors for taxoid biosynthesis in plant cells.
