Effect of sodium nitroprusside on growth and terpenoid indole alkaloid production in Catharanthus roseus hairy root cultures.

Nitric oxide (NO) is known as a signaling molecule involved in elicitor-induced defense responses of plants. Sodium nitroprusside (SNP), a donor of NO, stimulates catharanthine formation in Catharanthus roseus cells.1 Two important terpenoid indole alkaloids produced in small quantities within C. roseus are vinblastine and vincristine which are being used clinically as anticancer drugs. We are interested in engineering C. roseus hairy roots to increase the production of the TIAs. The present work investigates the effects of treating different concentrations of SNP to the hairy root cultures from line LBE-6-1. The alkaloid concentrations were analyzed 9, 14, 17, 20, 23, 26, and 30 days after treatment of SNP on day 0. We also studied the transient effects of SNP treatment during the exponential phase in C. roseus hairy roots. Analysis of the results showed that treatment of 0.1-mM SNP did not affect the growth of hairy roots, whereas 1-mM SNP suppressed the growth significantly, and 10-mM SNP almost completely inhibited the growth of hairy roots. 0.1-mM SNP treatment on day 0 caused a significant increase in the concentration of serpentine, catharanthine, ajmalicine, lochnericine and tabersonine production. SNP treatment on day 12 stimulated the formation of serpentine, catharanthine, ajmalicine, hörhammericine, lochnericine and tabersonine by day 21. After the initial stimulation, serpentine, horhammericine and lochnericine concentrations returned to the basal level by day 28. Treatment of 0.1-mM SNP on day 0 caused significant decrease in the mRNA levels for TDC, ASA, STR, ORCA3, ZCT1, and Crgbf1 on day 23. Treating 0.1-mM SNP on day 12 caused decreases in the expression levels of STR, ORCA3, ZCT1, and Crgbf1 on day 21 and day 28. Compared with day 28, the mRNA transcript of ZCT1 on day 21 is about twofold higher. Expression levels of G10H increased significantly.