The ORCA2 transcription factor plays a key role in regulation of the terpenoid indole alkaloid pathway

Chun Yao Li,Guy W Sander,Jacqueline V Shanks,Le Zhao,Alex L Leopold,Susan I Gibson

doi:10.1186/1471-2229-13-155

Abstract

BackgroundThe terpenoid indole alkaloid (TIA) pathway leads to the production of pharmaceutically important drugs, such as the anticancer compounds vinblastine and vincristine. Unfortunately, these drugs are produced in trace amounts, causing them to be very costly. To increase production of these drugs, an improved understanding of the TIA regulatory pathway is needed. Towards this end, transgenic Catharanthus roseus hairy roots that overexpress the ORCA2 TIA transcriptional activator were generated and characterized.ResultsTranscriptional profiling experiments revealed that overexpression of ORCA2 results in altered expression of key genes from the indole and terpenoid pathways, which produce precursors for the TIA pathway, and from the TIA pathway itself. In addition, metabolite-profiling experiments revealed that overexpression of ORCA2 significantly affects the levels of several TIA metabolites. ORCA2 overexpression also causes significant increases in transcript levels of several TIA regulators, including TIA transcriptional repressors.ConclusionsResults presented here indicate that ORCA2 plays a critical role in regulation of TIA metabolism. ORCA2 regulates expression of key genes from both feeder pathways, as well as the genes (STR and SGD) encoding the enzymes that catalyze the first two steps in TIA biosynthesis. ORCA2 may play an especially important role in regulation of the downstream branches of the TIA pathway, as it regulates four out of five genes characterized from this part of the pathway. Regulation of TIA transcriptional repressors by ORCA2 may provide a mechanism whereby increases in TIA metabolite levels in response to external stimuli are transient and limited in magnitude.

Highlights

The terpenoid indole alkaloid (TIA) pathway leads to the production of pharmaceutically important drugs, such as the anticancer compounds vinblastine and vincristine
Generation of C. roseus transgenic hairy root lines expressing ORCA2 under the control of an ethanol-inducible promoter To date, only a few studies have characterized ORCA2 [18,31] and no published studies have determined which TIA genes, other than Strictosidine synthase (STR), are regulated by ORCA2 or how ORCA2 overexpression affects the levels of a broad group of TIA metabolites
In addition to regulating some of the TIA transcriptional activator genes, ORCA2 regulates the TIA transcriptional repressor genes ZCT1, ZCT2 and ZCT3, but not GBF1 or GBF2. These findings provide a possible explanation for how overexpression of the ORCA2 transcriptional activator gene may lead indirectly to decreased strictosidine β-D-glucosidase (SGD) and deacetylvindoline acetyltransferase (DAT) transcript levels by causing overexpression of the ZCT1, ZCT2 and ZCT3 TIA transcriptional repressors, one or more of which may turn down expression of SGD and DAT

Summary

Introduction

The terpenoid indole alkaloid (TIA) pathway leads to the production of pharmaceutically important drugs, such as the anticancer compounds vinblastine and vincristine. These drugs are produced in trace amounts, causing them to be very costly. To increase production of these drugs, an improved understanding of the TIA regulatory pathway is needed. Towards this end, transgenic Catharanthus roseus hairy roots that overexpress the ORCA2 TIA transcriptional activator were generated and characterized. Ajmalicine and serpentine are sometimes used as anti-hypertensive agents Most of these alkaloids are produced in extremely low amounts in planta, limiting the usage of these chemicals.

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