S105: Examining the complex transcriptional response of perturbing one gene in the terpenoid indole alkaloid pathway in Catharanthus roseus

Catharanthus roseus (Madagascar periwinkle) produces two clinically important chemotherapy drugs, vinblastine and vincristine.  These drugs are produced through the terpenoid indole alkaloid (TIA) pathway.  While researchers have been studying the TIA pathway genes and regulators for the past 40 years, there are still many aspects of the pathway that are unknown.  A previous study has shown that overexpressing a feedback insensitive anthranilate synthase (AS) under the control of an inducible promoter in C. roseus hairy roots increases the levels of some TIA metabolites while decreasing the levels of other TIA metabolites.

Although the genetic modification is targeted to AS, we hypothesized that expression of other genes in the TIA pathway would be altered.  In order to determine the transcriptional response caused by the overexpression of AS, we utilized RT-qPCR to analyze the transcriptional response of all known TIA genes and regulators after 12, 24, and 48 hours of induction (i.e. increased expression of AS).  The results indicate that overexpression of AS caused an increase in known terpenoid pathway gene transcripts, indole pathway gene transcripts, and some TIA pathway gene transcripts.  Notably two TIA gene transcripts were significantly downregulated under the same conditions.  Similarly transcripts of TIA regulators were differentially expressed when AS was overexpressed.  These results will be discussed in greater detail along with their implications to future metabolic engineering efforts in plants.