Importance of Hormonal Elicitors in Inducing Morphine Biosynthesis in the Cell Culture of (Papaver bracteatum Lindl.)

Authors
M. Behzadirad 1
M. R. Naghavi 2
A. A. Shahnejat Bushehri 1
1 Agronomy and Plant Breeding Department, Agricultural College, University of Tehran, Karaj, Islamic Republic of Iran.
2 Department of Agronomy and Plant Breeding, College of Agriculture, University of Tehran, Karaj, Islamic Republic of Iran.
Abstract
Plant cells have enough capacity to produce many of secondary metabolites, similar to the whole plants. Elicitation is one of the most significant methods to increase the synthesis of secondary metabolites in the medicinal plants. The purpose of this study was to investigate the effect of three different hormones on alkaloids production in suspension culture of Papaver bracteatum Lindl., in order to identify the relationship between the alkaloid biosynthesis and gene expression. Inducible factors initiate BenzylIsoquinoline Alkaloids (BIAs) biosynthesis in Opium poppy. The current study investigated the accumulation of alkaloids content and Tyrosine/DopadeCarboxilase (TYDC), Berberine Bridge Enzyme (BBE), Salutardinol Acetyl Transferase (SAT), and Codeinone Reductase (COR) gene transcripts in suspension culture of P. bracteatum. Indole-3-Acetic Acid (IAA), Indole Butyric Acid (IBA) and Gibberellic Acid (GA) were used as hormonal elicitors in the suspension cultures with three different doses and two timings along with the control. This research showed the induction of morphine alkaloid in the suspension culture of P. bracteatum. Elicitation by 20 mg L-1 concentration of IAA after 48 h indicated significant increase in morphine amount. Comparison among genes revealed that the expression levels of COR dramatically increased while TYDC, BBE and SAT had no significant difference compared to the control. After elicitation by IAA, IBA, and GA, the highest levels of morphine were measured as 243.2, 207.2, and 178.1 mg g-1, respectively. The results demonstrated that timing had a significant effect on the hormonal elicitation: 48 h treatment could induce more morphine alkaloids compared to 24 hours treatments.

Keywords

Subjects

1. Alcantara, J., Bird, D. A., Franceschi, V. R. and Facchini, P.J. 2005. Sanguinarine Biosynthesis is Associated with the Endoplasmic Reticulum in Cultured Opium Poppy Cells after Elicitor Treatment. Plant Physiol., 138(1): 173-183.
2. Balažová, A., Blanáriková, V., Bilka, F. and Bilková, A. 2008. Effect of Combined Biotic and Abiotic Elicitor on the Sanguinarine Formation in Cell Suspension Cultures of Eschscholtzia Californica Cham. Acta Facult. Pharm., 55: 58-63.
3. Bohm, H. 1967. On the Papaver bracteatum Lindl. Characteristic Changes of Alkaloid Spectrum during Plant Development. Planta Med., 15: 215–220.
4. Brochmann‐Hanssen, E. and Wunderly, S. W. 1978. Biosynthesis of Morphine Alkaloids in Papaver bracteatum Lindl. J. Pharm. Sci., 67: 103-106.
5. Cline, S. D. and Coscia, C. J. 1988. Stimulation of Sanguinarine Production by Combined Fungal Elicitation and Hormonal Deprivation in Cell Suspension. Plant Physiol., 86: 0161-0165.
6. Cline, S. D. and Coscia, C. J. 1988. Stimulation of Sanguinarine Production by Combined Fungal Elicitation and Hormonal Deprivation in Cell Suspension Cultures of Papaver bracteatum. Plant Physiol., 86(1): 161-165.
7. Day, K.B., Draper, J. and Smith, H. 1986. Plant Regeneration and Thebaine Content of Plants Derived from Callus Culture of Papaverbracteatum. Plant Cell Reports, 5: 471–4.
8. Doke, N. 1983. Involvement of Superoxide Anion Generation in the Hypersensitive Response of Potato Tuber Tissues to Infection with an Incompatible Race of Phytophthorainfestans and to the Hyphal Wall Components. Physiol. Plant Pathol. 23(3): 345–357.
9. Farjaminezhad, R., Zare, N., Asghari-Zakaria, R. and Farjaminezhad, M. 2013. Establishment and Optimization of Cell Growth in Suspension Culture of Papaver bracteatum: A Biotechnology Approach for Thebaine Production. Turk. J. Biol., 37: 689-697.
10. Facchini, P. J., Penzes, C., Johnson, A. G., and Bull, D. 1996. Molecular Characterization of Berberine Bridge Enzyme Genes from Opium Poppy. Plant physiol., 112(4): 1669-1677.
11. Grothe, T., Lenz, R. and Kutchan, T. M. 2001. Molecular Characterization of the Salutaridinol 7-O-Acetyltransferase Involved in Morphine Biosynthesis in Opium Poppy Papaver somniferum. J. Biol. Chem., 276: 30717-30723.
12. Hodges, C., Horn, J. S. and Rapoport, H. 1977. Morphinan Alkaloids in Papaver bracteatum: Biosynthesis and Fate. Phytochemistry, 16: 1939-1942.
13. Hook, I., Sheridan, H. and Wilson, G. 1988. Alkaloids of Cell Cultures Derived from Strains of Papaver bracteatum. Phytochemistry, 27(7): 2137-2141.
14. Kamimura, S., Akutsu, M. and Nishikawa, M. 1976. Formation of Thebaine in the Suspension Culture of Papaver bracteatum. Agric. Biol. Chem., 40: 913-919.
15. Kamo, K. K., Chang, K. L., Lynn, M. E., and Hodges, T. K., 1987. Embryogenic Callus Formation from Maize Protoplasts. Planta, 172(2): 245-251.
16. Kunz, D. A., Reddy, G. S. and Vatvars, A. R. 1985. Identification of Transformation Products Arising from Bacterial Oxidation of Codeine by Streptomyces griseus. Appl. Environ. Microbiol., 50: 831-836.
17. Larkin, P.J., Miller, J. A., Allen, R.S., Chitty, J. A., Gerlach, W. L., Frick S., Kutchan T. M. and Fist, A. J. 2007. Increasing Morphinan Alkaloid Production by Over-Expressing Codeinone Reductase in Transgenic Papaver somniferum. Plant Biotechnol. J., 5: 26-37.
18. Namdeo, A. G. 2007. Plant Cell Elicitation for Production of Secondary Metabolites. Pharmacognosy Rev., 1: 69-79.
19. Nasiri, J., Naghavi, M. R., Alizadeh, H., Moghadam, M. R. 2016. Seasonal-based Temporal Changes Fluctuate Expression Patterns of TXS, DBAT, BAPT and DBTNBT Genes alongside Production of Associated Taxanes in Taxus baccata. Plant Cell Rep., 35(5):1103-1119.
20. Ranasinghe, S., Rogers, M. E., Hamilton, J. G., Bates, P. A. and Maingon, R. D. 2008. A Real-Time PCR Assay to Estimate Leishmania chagasi Load in Its Natural Sand Fly Vector Lutzomyia longipalpis. Trans. Royal Soc. Tropic. Med. Hygiene Sep., 102: 875-882.
21. Rostampour, S., Sohi, H. H., Jourabchi, E. and Ansari, E. 2009. Influence of Agrobacterium rhizogeneson Induction of Hairy Roots and Benzylisoquinoline Alkaloids Production in Persian Poppy (Papaver bracteatumLindl.): Preliminary Report. World J. Microb. Biot., 25: 1807-1814.
22. Rush, M. D., Kutchan, T. M. and Coscia, C. J. 1985. Correlation of Theappearance of Morphinan Alkaloids and Laticifer Cells in Germinating Papaver bracteatum Seedlings. Plant Cell Rep., 4: 237-240.
23. Samanani, N., Alcantara, J., Bourgault, R., Zulak, K. G. and Facchini, P. J. 2006. The Role of Phloem Sieve Elements and Laticifers in the Biosynthesis and Accumulation of Alkaloids in Opium poppy. Plant J., 47(4): 547-563.
24. Sharafi, A., Hashemi Sohi, H. , Mousavi, A., Azadi, P., Dehsara, B. and Hosseini Khalifani, B. 2013. Enhanced Morphinan Alkaloid Production in Hairy Root Cultures of Papaver bracteatum by Over-Expression of Salutaridinol b7-O-Acetyltransferase Gene via Agrobacterium rhizogenes Mediated Transformation. World J. Micro. Biol. Biotechnol., 29: 2125–2131.
25. Tisserat, B. and Berhow, M. 2009. Production of Pharmaceuticals from Papaver Cultivars in Vitro. Eng. Life Sci., 9: 190–196.
26. Unterlinner, B., Lenz, R., and Kutchan, T. M. 1999. Molecular Cloning and Functional Expression of Codeinone Reductase: the Penultimate Enzyme in Morphine Biosynthesis in the Opium Poppy Papaver somniferum. Plant J., 18(5): 465-475.
27. Zare, N., Farjaminezhad, R., Asghari-Zakaria, R. and Farjaminezhad, M. 2014. Enhanced Thebaine Production in Papaver bracteatum Cell Suspension Culture by Combination of Elicitation and Precursor Feeding. Nat. Prod. Res., 28: 711-717.
28. Zhang, L. and Xing, D. 2008. Methyl Jasmonate Induces Production of Reactive Oxygen Species and Alterations in Mitochondrial Dynamics that Precede Photosynthetic Dysfunction and Subsequent Cell Death. Plant Cell Physiol., 49: 1092-1111.
Journal of Agricultural Science and Technology
Volume 22, Issue 1
January and February 2020
Pages 261-270