Artemisinin, produced by the plant Artemisia annua, is a sesquiterpene anti-malarial therapeutic. Due to the medicinal relevance of this plant product, there is significant interest in understanding how the biosynthetic pathway is regulated at several key steps. The objective of this study is to examine several factors known to influence artemisinin yields to determine if those effects are occurring at the transcriptional level of the biosynthetic pathway. Artemisinin content has been shown to increase as the plant shifts from vegetative growth to reproductive, flowering growth. To test whether there is a corresponding increase in terpenoid gene expression during the shift to reproductive growth, levels of mRNA of terpenoid genes were measured during flowering budding and full flowering and compared to those measured during vegetative growth. Results indicate that in response to the photoperiod signal to shift to reproductive growth, early cytosolic pathway genes were highly upregulated, while there was no change in early plastidic pathway genes. Late pathway genes specific to artemisinin synthesis were upregulated >6-fold. Furthermore, glucose has also been shown to stimulate artemisinin production compared to sucrose. To test whether glucose is acting as signal to increase terpenoid gene expression, levels of mRNA of terpenoid genes were measured in glucose- and fructose-treated seedlings and compared to those in sucrose-treated seedlings. Results indicate that in response to treatment with glucose, compared with sucrose, early pathway genes in both compartments were initially upregulated. Transcript levels subsequently decreased to levels similar to those in sucrose-treated seedlings. ADS was upregulated by glucose, compared with sucrose, reaching a peak at day 7. Finally, coordinate control of sterol and sesquiterpene synthesis at a critical branch-point in the terpenoid biosynthetic pathway has been demonstrated. To test whether amorpha-4,11-diene synthase (ADS) and squalene sythase (SQS) are coordinately regulated, levels of mRNA of those two genes were measured and compared in both experimental conditions. Results indicate that under the conditions used in this study, ADS and SQS did not show coordinate regulation. This study was the first to demonstrate that: 1. terpenoid genes relating to artemisinin biosynthesis are regulated at the level of transcript accumulation as the plant shifts from vegetative to reproductive growth; 2. glucose is acting as a signal in artemisinin biosynthesis by upregulating transcript levels for several terpenoid genes.
| Identifer | oai:union.ndltd.org:wpi.edu/oai:digitalcommons.wpi.edu:etd-theses-1414 |
| Date | 28 April 2008 |
| Creators | Vail, Daniel Robert |
| Contributors | Pamela J. Weathers, Advisor, Reeta Prusty Rao, Committee Member, , Kristin K. Wobbe |
| Publisher | Digital WPI |
| Source Sets | Worcester Polytechnic Institute |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Masters Theses (All Theses, All Years) |
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