In collaboration with Scientific Association of Iranian Medicinal Plants

Document Type : Research Paper

Authors

1 M.Sc. student, Department of Genetics and Plant Breeding, Ahvaz Branch, Islamic Azad University, Ahvaz, Iran

2 Faculty member, Department of Genetics and Plant Breeding, Ahvaz Branch, Islamic Azad University, Ahvaz, Iran

3 Faculty Member, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran

Abstract

Identification of the genes biosynthesizing medicinal plants-specific metabolites is now performed with great speed and accuracy using new transcriptome study technologies such as RNA sequencing. The present study was carried out to find the specific genes in the biosynthetic pathway of triterpenes and sesquiterpenes in the fruit tissue of colocynth (Citrullus colocynthis L.). After RNA extraction from the tissue of colocynth fruits harvested from Andimeshk region in Khuzestan province in 2017, RNA sequencing technique was performed using the Illumina HiSeq2500 platform. The bioinformatics steps including de novo assembly, using the Evidential-gene software, and functional annotation, using the KAAS database, were performed. In the KAAS database, 17359 unigenes were annotated in 134 plant pathways. Among the different and important secondary metabolites pathways in the fruit tissue of colocynth, 39 unigenes and 8 orthologous genes were assigned to the triterpenes and sesquiterpenes gene pathways. Transcriptome analysis of this medicinal plant with the aim of identifying the genes of secondary metabolites biosynthetic pathways underlies various research and practical aspects such as biosynthetic pathway engineering of herbal medicines.

Keywords

- Abo, K., Fred-Jaiyesimi, A. and Jaiyesimi, A., 2008. Ethnobotanical studies of medicinal plants used in the management of diabetes mellitus in South Western Nigeria. Journal of Ethnopharmacology, 115(1): 67-71.
- Amiripour, M., Sadat Nouri, S.A., Shariati, V. and Soltani Howyzeh, M., 2018. Identification of terpenoid backbone biosynthetic pathway genes in Ajowan (Trachyspermum ammi L.) by RNA-Seq. Modern Genetic Journal, 13(1): 133-141.
- Bains, S., Thakur, V., Kaur, J., Singh, K. and Kaur, R., 2018. Elucidating genes involved in sesquiterpenoid and flavonoid biosynthetic pathways in Saussurea lappa by de novo leaf transcriptome analysis. Genomics, 116(6): 1474-1482.
- Bhambhani, S., Lakhwani, D., Gupta, P., Pandey, A., Dhar, Y.V., Bag, S.K., Asif, M.H. and Trivedi, P.K., 2017. Transcriptome and metabolite analyses in Azadirachta indica: identification of genes involved in biosynthesis of bioactive triterpenoids. Scientific Reports, 7(5043): 1-12.
- Boutanaev, A.M., Moses, T., Zi, J., Nelson, D.R., Mugford, S.T., Peters, R.J. and Osbourn, A., 2015. Investigation of terpene diversification across multiple sequenced plant genomes. Proceedings of the National Academy of Sciences, 112(1): E81-E88.
- Celedon, J.M., Chiang, A., Yuen, M.M., Diaz-Chavez, M.L., Madilao, L.L., Finnegan, P.M., Barbour, E.L. and Bohlmann, J., 2016. Heartwood‐specific transcriptome and metabolite signatures of tropical sandalwood (Santalum album) reveal the final step of (Z)‐santalol fragrance biosynthesis. The Plant Journal, 86(4): 289-299.
- Chen, Q., Ma, C., Qian, J., Lan, X., Chao, N., Sun, J. and Wu, Y., 2016. Transcriptome sequencing of Gynostemma pentaphyllum to identify genes and enzymes involved in Triterpenoid biosynthesis. International Journal of Genomics, 2016: 7840914.
- Cuong, D.M., Jeon, J., Morgan, A.M., Kim, C., Kim, J.K., Lee, S.Y. and Park, S.U., 2017. Accumulation of charantin and expression of triterpenoid biosynthesis genes in bitter melon (Momordica charantia). Journal of agricultural and food chemistry, 65(33): 7240-7249.
- Dorafshan, M., Soltani Howyzeh, M. and Shariati, V., 2019. Identification of terpenoid backbone biosynthetic pathway genes in fruit of Citrullus colocynthis medical plant by RNA sequencing. Iranian Journal of Medicinal and Aromatic Plants Research, 35(4): 691-702.
- Gurudeeban, S. and Ramanathan, T., 2010. Antidiabetic effect of Citrullus colocynthis in alloxan-induced diabetic rats. Inventi Rapid: Ethnopharmacology, 1: 12.
- Han, X.J., Wang, Y.D., Chen, Y.C., Lin, L.Y. and Wu, Q.K., 2013. Transcriptome sequencing and expression analysis of terpenoid biosynthesis genes in Litsea cubeba. PLOS ONE, 8(10): e76890.
- Javadzadeh, H.R., Davoudi, A., Davoudi, F., Valizadegan, G., Goodarzi, H., Mahmoodi, S., Ghane, M.R. and Faraji, M., 2013. Citrullus colocynthis as the cause of acute rectorrhagia. Case Reports in Emergency Medicine, 5p.
- Jiang, Z., Kempinski, C., Bush, C.J., Nybo, S.E. and Chappell, J., 2016. Engineering triterpene and methylated triterpene production in plants provides biochemical and physiological insights into terpene metabolism. Plant Physiology, 170(2): 702-716.
- Langenheim, J.H., 1994. Higher plant terpenoids: a phytocentric overview of their ecological roles. Journal of Chemical Ecology, 20(60): 1223-1280.
- Lee, H.A., Kim, S., Kim, S. and Choi, D., 2017. Expansion of sesquiterpene biosynthetic gene clusters in pepper confers nonhost resistance to the Irish potato famine pathogen. New Phytologist, 215(3): 1132-1143.
- Luo, H., Sun, C., Sun, Y., Wu, Q., Li, Y., Song, J., Niu, Y., Cheng, X., Xu, H. and Li, C., 2011. Analysis of the transcriptome of Panax notoginseng root uncovers putative triterpene saponin-biosynthetic genes and genetic markers. BMC Genomics, 12(5): S5.
- Miettinen, K., Inigo, S., Kreft, L., Pollier, J., De, Bo. C., Botzki, A., Coppens, F., Bak, S. and Goossens, A., 2017a. The TriForC database: a comprehensive up-to-date resource of plant triterpene biosynthesis. Nucleic Acids Research, 46(1): D586-D594.
- Miettinen, K., Pollier, J., Buyst, D., Arendt, P., Csuk, R., Sommerwerk, S., Moses, T., Mertens, J., Sonawane, P.D. and Pauwels, L., 2017b. The ancient CYP716 family is a major contributor to the diversification of eudicot triterpenoid biosynthesis. Nature Communications, 8: 14153.
- Moses, T., Papadopoulou, K.K. and Osbourn, A., 2014. Metabolic and functional diversity of saponins, biosynthetic intermediates and semi-synthetic derivatives. Critical Reviews in Biochemistry and Molecular Biology, 49(6): 439-462.
- Moses, T., Pollier, J., Shen, Q., Soetaert, S., Reed, J., Erffelinck, M.L., Van, Nieuwerburgh, F.C., Bossche,. R.V., Osbourn, A. and Thevelein, J.M., 2015. OSC2 and CYP716A14v2 catalyze the biosynthesis of triterpenoids for the cuticle of aerial organs of Artemisia annua. The Plant Cell, 27(1): 286-301.
- Padilla-Gonzalez, G.F., dos Santos, F.A. and Da Costa, F.B., 2016. Sesquiterpene lactones: more than protective plant compounds with high toxicity. Critical Reviews in Plant Sciences, 35(1): 18-37.
- Rabbani, B., Nakaoka, H., Akhondzadeh, S., Tekin, M. and Mahdieh, N., 2016. Next generation sequencing: implications in personalized medicine and pharmacogenomics. Molecular BioSystems, 12(6): 1818-1830.
- Rai, A., Yamazaki, M., Takahashi, H., Nakamura, M., Kojoma, M., Suzuki, H. and Saito, K., 2016. RNA-seq transcriptome analysis of Panax japonicus, and its comparison with other Panax species to identify potential genes involved in the saponins biosynthesis. Frontiers in Plant Science, 7: 481.
- Seki, H., Tamura, K. and Muranaka, T., 2015. P450s and UGTs: key players in the structural diversity of triterpenoid saponins. Plant and Cell Physiology, 56(8): 1463-1471.
- Si, Y., Dane, F., Rashotte, A., Kang, K. and Singh, N.K., 2010. Cloning and expression analysis of the Ccrboh gene encoding respiratory burst oxidase in Citrullus colocynthis and grafting onto Citrullus lanatus (watermelon). Journal of Experimental Botany, 61(6): 1635-1642.
- Soltani Howyzeh, M., Sadat Nouri, S.A., Shariati, V. and Amiripour, M., 2018. Comparative transcriptome analysis to identify putative genes involved in thymol biosynthesis pathway in medicinal plant Trachyspermum ammi L. Scientific Reports, 8(1): 13405.
- Sun, H., Li, F., Xu, Z., Sun, M., Cong, H., Qiao, F. and Zhong, X., 2017. De novo leaf and root transcriptome analysis to identify putative genes involved in triterpenoid saponins biosynthesis in Hedera helix L. PLOS ONE, 12(8): e0182243.
- Tang, Q., Ma, X., Mo, C., Wilson, I.W., Song, C., Zhao, H., Yang, Y., Fu, W. and Qiu, D., 2011. An efficient approach to finding Siraitia grosvenorii triterpene biosynthetic genes by RNA-seq and digital gene expression analysis. BMC Genomics, 12(1): 343.
- Tao, T., Liu, X., Chang, J., Xu, F. and Yin, Y., 2016. Cloning and characterisation of the gene encoding acetyl-coa c-acetyltransferase in Matricaria chamomilla. Journal of Pharmaceutical, Chemical and Biological, 4(3): 386-393.
- Thimmappa, R., Geisler, K., Louveau, T., O'Maille, P. and Osbourn, A., 2014. Triterpene biosynthesis in plants. Annual Review of Plant Biology, 65: 225-257.
- Tian, X., Ruan, J.X., Huang, J.Q., Yang, C.Q., Fang, X., Chen, Z.W., Hong, H., Wang, L.J., Mao, Y.B. and Lu, S., 2018. Characterization of gossypol biosynthetic pathway. Proceedings of the National Academy of Sciences, 115(23): E5410-E5418.
- Uma, C. and Sekar, K., 2014. Phytochemical analysis of a folklore medicinal plant Citrullus colocynthis L. (bitter apple). Journal of Pharmacognosy and Phytochemistry, 2(6): 195-202.
- Upadhyay, B., Roy, S. and Kumar, A., 2007. Traditional uses of medicinal plants among the rural communities of Churu district in the Thar Desert, India. Journal of Ethnopharmacology, 113(3): 387-399.
- Wang, Z., Hu, H., Goertzen, L.R., McElroy, J.S. and Dane, F., 2014. Analysis of the Citrullus colocynthis transcriptome during water deficit stress. PLoS One, 9(8): e104657.
- Wu, D., Austin, R.S., Zhou, S. and Brown, D., 2013. The root transcriptome for North American ginseng assembled and profiled across seasonal development. BMC Genomics, 14(1): 564.
- Zhan, C., Li, X., Zhao, Z., Yang, T., Wang, X., Luo, B., Zhang, Q., Hu, Y. and Hu, X., 2016. Comprehensive analysis of the triterpenoid saponins biosynthetic pathway in Anemone flaccida by transcriptome and proteome profiling. Frontiers in Plant Science, 7: 1094.
- Zhang, W., Tao, T., Liu, X., Xu, F., Chang, J. and Liao, Y., 2018. De novo assembly and comparative transcriptome analysis: novel insights into sesquiterpenoid biosynthesis in Matricaria chamomilla L. Acta Physiologiae Plantarum, 40(7): 129.
- Zheng, X., Xu, H., Ma, X., Zhan, R. and Chen, W., 2014. Triterpenoid saponin biosynthetic pathway profiling and candidate gene mining of the Ilex asprella root using RNA-Seq. International Journal of Molecular Sciences, 15(4): 5970-5987.