Abstract
The spatial and temporal expression of dbat and dbtnbt genes involved in the later steps of paclitaxel biosynthesis in relation to baccatin III and paclitaxel accumulation in Taxus baccata L. germinating embryos and seedlings was investigated. The steady-state of mRNA transcripts was measured by quantitative real time polymerase chain reaction (qRT-PCR), the content of taxanes was determined by HPLC. The spatial distribution of the metabolites was found to be in accordance with the transcript level of the respective genes. Higher content of mRNA transcripts in shoots of yew seedlings responded to higher content of taxanes in stems and needles. The highest increase in the transcript level of both genes was observed 8 d after placing the embryos on germination medium, before elongation of embryonic axis and emergence of the radicle. The pattern of temporal dbat expression was in line with the expression of the dbtnbt gene. The temporal distribution of the precursor baccatin III correlated well with paclitaxel, the final product of the pathway.
This is a preview of subscription content, access via your institution.
Abbreviations
- DBAT:
-
10-deacetylbaccatin-III-10β-O-acetyltransferase
- DBTNBT:
-
3′-N-debenzoyl-2′-deoxytaxol-N-benzoyltransferase
- REU:
-
relative expression units
- RT-PCR:
-
quantitative real time polymerase chain reaction
References
Bonfill, M., Bentebibel, S., Moyano, E., Palazón, J., Cusidó, R.M., Eibl, R., Pinol, M.T.: Paclitaxel and baccatin III production induced by methyl jasmonate in free and immobilized cells of Taxus baccata. — Biol. Plant. 51: 647–652, 2007.
Bruňáková, K., Babincová, Z., Takáč, M., Čellárová, E.: Selection of callus cultures of Taxus baccata L. as a potential source of paclitaxel production. — Engn. Life Sci. 5: 465–469, 2004.
Canas, R.A., Torre, F., Canovas, F.M., Canton, F.R.: High levels of asparagin synthetase in hypocotyls of pine seedlings suggest a role of the enzyme in re-allocation of seed-stored nitrogen. — Planta 224: 83–95, 2006.
Chang, S.H., Ho, C.K., Chen, Z.Z., Tsay, J.Y.: Micropropagation of Taxus mairei from mature trees. — Plant Cell. Rep. 20: 496–502, 2001.
Chee, P.P.: Organogenesis in Taxus brevifolia tissue cultures. — Plant Cell Rep. 14: 560–565, 1995.
Croteau, R., Ketchum, R.E.B., Long, R.M., Kaspera, R., Wildung, M.R.: Taxol biosynthesis and molecular genetics. — Phytochem. Rev. 5: 75–97, 2006.
Fett-Neto, A.G., DiCosmo, F.: Distribution and amounts of taxol in different shoot parts of Taxus cuspidata. — Planta Med. 58: 464–466, 1992.
Fett-Neto, A.G., DiCosmo, F.: Taxol and taxane production by cell culture. — In: Meyers, R.A. (ed.): Encyclopedia of Molecular Biology and Molecular Medicine. Vol. 6. Pp. 10–17. VCH Verlagsgesellschaft, Weinheim 1997.
Flores, H.E., Sgrignoli, P.J.: In vitro culture and precocious germination of Taxus embryos. — In Vitro cell. dev. Biol. Plant 27: 139–142, 1991.
Gallardo, K., Job, C., Groot, S.P.C., Puype, M., Demol, H., Vandekerckhove, J., Job, D.: Proteomic analysis of Arabidopsis seed germination and priming. — Plant Physiol. 126: 835–848, 2001.
Goncalves, S., Cairney, J., Maroco, J., Oliveira, M.M., Miguel, C.: Evaluation of control transcripts in real-time RT PCR expression analysis during maritime pine embryogenesis. — Planta 222: 556–563, 2005.
Guo, B.H., Kai, G.Y., Jin, H.B., Tang, K.X.: Taxol synthesis. — Afr. J. Biotechnol. 5: 15–20, 2006.
Hefner, J., Ketchum, R.E.B., Croteau, R.: Cloning and functional expression of a cDNA encoding geranylgeranyl diphosphate synthase from Taxus canadensis and assessment of the role of this prenyltransferase in cells induced for Taxol production. — Arch. Biochem. Biophys. 360: 62–74, 1998.
Horowitz, S.B., Lothsteia, L., Manfredi, J.J., Mellado, W., Parness, J., Roy, S.N., Schiff, P.B., Sorbara, L., Zeheb, R.: Taxol: mechanism of action and resistance. — Ann. N.Y. Acad. Sci. 466: 733–743, 1986.
Kai, G., Miao, Z., Qiu, C., Zhang, L., Gong, Y., Zhao, D., Liu, D., Sun, X., Tang, K.: Molecular cloning and characterization of a taxadienol acetyl transferase cDNA from Taxus × media. — Plant Sci. 167: 759–764, 2004.
Kai, G., Miao, Z., Zhang, L., Zhao, D., Liao, Z., Sun, X., Zhao, L., Tang, K.: Molecular cloning and expression analyses of a new gene encoding 3-hydroxy-3-methylglutaryl-CoA synthase from Taxus × media. — Biol. Plant. 50: 359–366, 2006.
Kai, G., Zhao, L., Zhang, L., Li, Z., Guo, B., Zhao, D., Sun, X., Miao, Z., Tang, K.: Characterization and expression profile analysis of a new cDNA encoding taxadiene synthase from Taxus media. — J. Biochem. mol. Biol. 38: 668–675, 2005.
Katkovčinová, Z., Lázárová, M., Bruňáková, K., Košuth, J., Čellárová, E.: Expression of dbat and dbtnbt genes involved in paclitaxel biosynthesis during growth cycle of Taxus baccata L. callus cultures. — Z. Naturforsch. 63c: 721–730, 2008
Liao, Z., Tan, Q., Chai, Y., Zuo, K, Chen, M., Gong, Y., Wang, P., Pi, Y., Tan, F., Sun, X., Tang, K.: Cloning and characterisation of the gene encoding HMG-CoA reductase from Taxus media and its functional identification in yeast. — Funct. Plant Biol. 31: 73–81, 2004.
McCaskill, D., Croteau, R.: Isopentenyl diphosphate is the terminal product of the deoxyxylulose-5-phosphate pathway for terpenoid biosynthesis in plants. — Tetrahedron Lett. 40: 653–656, 1999.
Nims, E., Dubois, C.P., Roberts, S.C., Walker, E.L.: Expression profiling of genes involved in paclitaxel biosynthesis for targeted metabolic engineering. — Metab. Engn. 8: 385–394, 2006.
Schiff, P.B., Fant, J., Horwitz, S.B.: Promotion of microtubule assembly in vitro by taxol. — Nature 277: 665–667, 1979.
Škrlep, K., Bergant, M., De Winter, G.M., Bohanec, B., Žel, J., Verpoorte, R., Van Iren, F., Camloh, M.: Cryopreservation of cell suspension cultures of Taxus × media and Taxus floridana. — Biol. Plant. 52: 329–333, 2008.
Tranbarger, T.J., Misra, S.: The molecular characterization of a set of cDNAs differentially expressed during Douglas-fir germination and early seedling development. — Physiol. Plant. 95: 456–464, 1995.
Turner, G., Gersherzon, J., Nielson, E.E.: Limonene synthase, the enzyme responsible for monoterpene biosynthesis in peppermint, is localized to leucoplasts of oil gland secretory cells. — Plant Physiol. 120: 879–886, 1999.
Wickremesinhe, E.R.M., Arteca, R.: Roots of hydroponically grown Taxus plants as a source of taxol and related taxanes. — Plant Sci. 101: 125–135, 1994.
Wildung, M.R., Croteau, R.: A cDNA clone for taxadiene synthase, the diterpene cyclase that catalyses the commited step of taxol biosynthesis. — J. biol. Chem. 271: 9201–9204, 1996.
Wu, Z.L., Yuan, Y.J., Liu, J.X., Xuan, H.Y., Hu, Z.D., Sun, A.C., Hu, C.X.: Study on enhanced production of taxol from Taxus chinensis var. mairei in biphasic-liquid culture. — Acta bot. sin. 41: 1108–1113, 1999.
Zhiri, A., Jaziri, M., Homes, J., Vanhaelen, M., Shimomura, K.: Factors affecting the in vitro rapid germination of Taxus embryos and the evaluation of taxol content in the plantlets. — Plant Cell Tissue Organ Cult. 39: 261–263, 1994.
Acknowledgements
This work was supported by grant of the Scientific Grant Agency (VEGA) of the Ministry of Education of the Slovak Republic, No. 1/1273/04. Authors would like to thank Mrs. Hana Michalčová for her technical assistance at taxane isolation and quantification.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Bruňáková, K., Košuth, J., Katkovčinová, Z. et al. Expression of two genes of paclitaxel biosynthetic pathway during germination of Taxus baccata zygotic embryos. Biol Plant 54, 515–519 (2010). https://doi.org/10.1007/s10535-010-0090-3
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10535-010-0090-3
Additional key words
- dbat, dbtnbt
- qRT-PCR
- yew