Saffron Agronomy and Technology

Saffron Agronomy and Technology

Identification and sequencing of CCD4a and CCD4b genes in wild saffron

Document Type : Research Paper

Authors
1 Associate Professor, Agronomy and Plant Breeding Department, Faculty of Agriculture, Ilam University, Ilam, Iran
2 Associate Professor, Department of plant production, University of Torbat Heydarieh, Torbat Heydarieh, Iran
3 Saffron institute, University of Torbat Heydarieh, Torbat Heydarieh, Iran
4 Ph. D. of Plant Breeding -Molecular genetics and genetic engineering,Agronomy and Plant Breeding Department, Faculty of Agriculture, Ilam University, Ilam, Iran
Abstract
Apocarotenoid compounds cause various communication functions in plants. Apocarotenoids are the result of enzymatic cleavage of carotenoids catalyzed by carotenoid dioxygenases (CCD). The CCD4 family is the largest family of plant CCDs. In this research, genomic DNA was extracted from fresh leaves of wild saffron Crocus haussknechtii BOISS and used as a template for amplification of CCD4a and CCD4b genes in PCR reaction. PCR products were sequenced after purification. Then, the gene sequence was compared with the saffron gene available in the gene bank and a phylogenetic tree related to their sequence was drawn. In addition, the amino acids of proteins were compared and the spatial structure of the protein of these two genes was drawn. The results showed that the number of nucleotides in CCD4a gene is equal to 2402 and in CCD4b gene is 2373 kb. The dendrogram related to the sequence of CCD4a gene showed that the species were divided into 5 separate groups based on the similarity in the sequence of nucleic acids, and the highest similarity was related to the CCD4a gene of Iranian agricultural saffron with a rate of 99.82. In addition, the dendrogram diagram for the CCD4b gene was divided into three groups, with the lowest distance between this gene and the genes of saffron species. By defining the exon regions, the protein sequence of CCD4a and CCD4b genes was determined, and their number of amino acids was equal to 577 and 567, respectively. The findings of this research can provide valuable information regarding the behavior and reaction of CCD4a and CCD4b enzymes in the synthesis of C. haussknechtii apocarotenoids and can be used in gene transfer programs from wild saffron to saffron be a useful crop.
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Ahrazem, O., Argandoña, J., Fiore, A., Aguado, C., Luján, R., Rubio-Moraga, Á., Marro, M., Araujo-Andrade, C., Loza-Alvarez, P., Diretto, G., & Gomez-Gomez, L. (2018). Transcriptome analysis in tissue sectors with contrasting crocins accumulation provides novel insights into apocarotenoid biosynthesis and regulation during chromoplast biogenesis. Scientific Reports8 (1), 2843-2859. https://doi.org/10.1038/s41598-018-21225-z.
Ahrazem, O., Argandoña, J., Fiore, A., Rujas, A., Rubio-Moraga, Á., Castillo, R., & Gómez-Gómez, L. (2019). Multi-species transcriptome analyses for the regulation of crocins biosynthesis in Crocus. BMC Genomics20, 1-15.
Ahrazem, O., Rubio-Moraga, A., Argandona-Picazo, J., Castillo, R., & Gómez-Gómez, L. (2016). Intron retention and rhythmic diel pattern regulation of carotenoid cleavage dioxygenase 2 during crocetin biosynthesis in saffron. Plant Molecular Biology91, 355-374. https://doi.org/10.1186/s12864-019-5666-5.
Al-Snafi, A. E. (2016). Nutritional value and pharmacological importance of citrus species grown in Iraq. IOSR Journal of Pharmacy6 (8), 76-108. https://doi.org/10.9790/3013-0680176108.
Anaeigoudari, A. (2022). Antidepressant and anti-nociceptive effects of Nigella sativa and its main constituent, thymoquinone: A literature review. Asian Pacific Journal of Tropical Biomedicine12 (12), 495-503. https://doi.org/10.4103/2221-1691.363875.
Beiki, A. H., Keify, F., & Mozafari, J. (2011). Rapid genomic DNA isolation from corm of Crocus species for genetic diversity analysis. Journal of Medicinal Plants Research5 (18), 4596-4600. https://doi.org/10.5897/JMPR.
Bukhari, S. I., Manzoor, M., & Dhar, M. K. (2018). A comprehensive review of the pharmacological potential of Crocus sativus and its bioactive apocarotenoids. Biomedicine & Pharmacotherapy98, 733-745. https://doi.org/10.1016/j.biopha.2017.12.090.
Cerdá-Bernad, D., Valero-Cases, E., Pastor, J. J., & Frutos, M. J. (2022). Saffron bioactives crocin, crocetin and safranal: Effect on oxidative stress and mechanisms of action. Critical Reviews in Food Science & Nutrition62 (12), 3232-3249. https://doi.org/10.1080/10408398.2020.1864279.
Cheng, B., Furtado, A., & Henry, R. J. (2017). Long-read sequencing of the coffee bean transcriptome reveals the diversity of full-length transcripts. Gigascience6 (11), 1-13. https://doi.org/10.1093/gigascience/gix086.
El Midaoui, A., Ghzaiel, I., Vervandier-Fasseur, D., Ksila, M., Zarrouk, A., Nury, T., Khallouki, F., El Hessni, A., Ibrahimi, S. O., Latruffe, N., & Lizard, G. (2022). Saffron (Crocus sativus L.): A source of nutrients for health and for the treatment of neuropsychiatric and age-related diseases. Nutrients14 (3), 597-617. https://doi.org/10.3390/nu14030597.
Eshaghi, M., & Rashidi-Monfared, S. (2024). Co-regulatory network analysis of the main secondary metabolite (SM) biosynthesis in Crocus sativus L. Scientific Reports14 (1), 15839- 1584. https://doi.org/10.1038/s41598-024-65870-z.
Ghahghaei, A., Bathaie, S. Z., Kheirkhah, H., & Bahraminejad, E. (2013). The protective effect of crocin on the amyloid fibril formation of Aβ42 peptide in vitro. Cellular & Molecular Biology Letters18, 328-339. https://doi.org/10.2478/s11658-013-0092-1.
Habibzadeh, M. J., Dorani, E., Ziaratnia, S. M., & Valizadeh, M. (2020). Cloning and bioinformatics investigation on CCD4a and CCD4b genes from Iranian saffron (Crocus sativus L.). Saffron Agronomy & Technology8 (2), 211-229 (In Persian with English Abstract). https://doi.org/10.22048/jsat.2023.403704.1492.
Hashimoto, H., Uragami, C., & Cogdell, R. J. (2016). Carotenoids and photosynthesis. Carotenoids in Nature: Biosynthesis, Regulation & Function, 111-139. https://doi.org/10.1007/978-3-319-39126-7_4.
Hou, X., Rivers, J., León, P., McQuinn, R. P., & Pogson, B. J. (2016). Synthesis and function of apocarotenoid signals in plants. Trends in Plant Science21 (9), 792-803. https://doi.org/10.1016/j.tplants.2016.06.001.
Izadpanah, F., Kalantari, S., Hassani, M. E., Naghavi, M. R., & Shokrpour, M. (2014). Variation in Saffron (Crocus sativus L.) accessions and Crocus wild species by RAPD analysis. Plant Systematics & Evolution300, 1941-1944.
Khakpour, A., Zolfaghari, M., & Sorkheh, K. (2019). Bioinformatics study and investigation of the expression pattern of several important genes involved in glycyrrhizin synthesis of Glycyrrhiza glabra L. in autumn and spring seasons. Plant Genetic Researches6 (1), 55-68. (In Persian with English Abstract). https://doi.org/10.29252/pgr.6.1.55.
Litt, A., & Irish, V. F. (2003). Duplication and diversification in the APETALA1/FRUITFULL floral homeotic gene lineage: implications for the evolution of floral development. Genetics165 (2), 821-833. https://doi.org/10.1093/genetics/165.2.821.
López, A., & Bonasora, M. G. (2017). Phylogeography, genetic diversity and population structure in a Patagonian endemic plant. AoB Plants, 9 (3), 1-12. https://doi.org/10.1093/aobpla/plx017.
Luo, D., Wang, T., Ye, M., Zhu, X., Cheng, Y., Zheng, Y., Xing, B & Shao, Q. (2023). Identification and characterization of Crocus sativus WRKY and its interacting MPK involved in crocins biosynthesis based on full-length transcriptome analysis. Industrial Crops & Products197, 1159-1165. https://doi.org/10.1016/j.indcrop.2023.116559.
Mosaviniya, M., Kikhavani, T., Tanzifi, M., Yaraki, M. T., Tajbakhsh, P., & Lajevardi, A. (2019). Facile green synthesis of silver nanoparticles using Crocus haussknechtii Bois bulb extract: Catalytic activity and antibacterial properties. Colloid & Interface Science Communications33, 100-111. https://doi.org/10.1016/j.colcom.2019.100211.
Mi, J., & Al-Babili, S. (2019). To color or to decolor: that is the question. Molecular Plant12 (9), 1173-1175. https://doi.org/10.1016/j.molp.2019.07.007. 
Moise, A. R., Al-Babili, S., & Wurtzel, E. T. (2014). Mechanistic aspects of carotenoid biosynthesis. Chemical Reviews114 (1), 164-193. https://doi.org/10.1021/cr400106y.
Molina, R. V., Valero, M., Navarro, Y., Guardiola, J. L., & Garcia-Luis, A. J. S. H. (2005). Temperature effects on flower formation in saffron (Crocus sativus L.). Scientia Horticulturae103 (3), 361-379. https://doi.org/10.1016/j.scienta.2004.06.005.
Moreno, J. C., Mi, J., Alagoz, Y., & Al‐Babili, S. (2021). Plant apocarotenoids: from retrograde signaling to interspecific communication. The Plant Journal105 (2), 351-375. https://doi.org/10.1111/tpj.15102.
Najari, G., Aazami, F., Taghi Mollaei, Y., & Fattahi, S., 2016. The morphological survey of wild saffron species in forests and rangeland of Ilam Province Ghasem. Forest Sterategical Approachment Journal, 1 (3), 46–53. )In Persian with English Abstract(. https://www.magiran.com/p1684320.
Pandita, D., Pandita, A., Wani, S. H., Abdelmohsen, S. A., Alyousef, H. A., Abdelbacki, A. M., Al-Yafrasi, M.A., Al-Mana, F. A., & Elansary, H. O. (2021). Crosstalk of multi-omics platforms with plants of therapeutic importance. Cells, 10 (6), 1296. https://doi.org/10.3390/cells10061296.
Qian, X., Sun, Y., Zhou, G., Yuan, Y., Li, J., Huang, H., & Li, L. (2019). Single-molecule real-time transcript sequencing identified flowering regulatory genes in Crocus sativusBMC Genomics20, 1-18. https://doi.org/10.1186/s12864-019-6200-5.
 Rodriguez-Concepcion, M., Avalos, J., Bonet, M. L., Boronat, A., Gomez-Gomez, L., Hornero-Mendez, D., & Zhu, C. (2018). A global perspective on carotenoids: Metabolism, biotechnology, and benefits for nutrition and health. Progress in Lipid Research70, 62-93. https://doi.org/10.1016/j.plipres.2018.04.004.
Rubio, A., Rambla, J. L., Santaella, M., Gomez, M. D., Orzaez, D., Granell, A., & Gomez-Gomez, L. (2008). Cytosolic and plastoglobule-targeted carotenoid dioxygenases from Crocus sativus are both involved in β-ionone release. Journal of Biological Chemistry283 (36), 24816-24825. https://doi.org/10.1074/jbc.M804000200.
Shahraki, H., Mahdinezhad, N., Fakheri, B., & Haddadi, F. (2020). Separation and identification of FEH1 gene in thorny artichoke plant (Cynara cardunculus) and its relative expression under the influence of abiotic stresses. Separation and identification of FEH1 gene in thorny artichoke plant (Cynara cardunculus) and its relative expression under the influence of abiotic stresses. Journal of Modern Genetics, 15 (2), 171-181 (In Persian with English Abstract). https://doi.org/20.1001.1.20084439.1399.15.2.9.2.
Su, C., Zhang, X., & Dubey, J. P. (2006). Genotyping of Toxoplasma gondii by multilocus PCR-RFLP markers: a high resolution and simple method for identification of parasites. International Journal for Parasitology, 36 (7), 841-848. https://doi.org/10.1016/j.ijpara.2006.03.003.
Tan, H., Chen, X., Liang, N., Chen, R., Chen, J., Hu, C & Zhang, L. (2019). Transcriptome analysis reveals novel enzymes for apo-carotenoid biosynthesis in saffron and allows construction of a pathway for crocetin synthesis in yeast. Journal of Experimental Botany70 (18), 4819-4834. https://doi.org/10.1093/jxb/erz211.
Tamura, K., Stecher, G., Peterson, D., Filipski, A., & Kumar, S. (2013). MEGA6: molecular evolutionary genetics analysis version 6.0. Molecular Biology & Evolution, 30 (12), 2725-2729. https://doi.org/10.1093/molbev/mst197.
Tiribuzi, R., Crispoltoni, L., Chiurchiù, V., Casella, A., Montecchiani, C., Del Pino, A. M., & Orlacchio, A. (2017). Trans-crocetin improves amyloid-β degradation in monocytes from Alzheimer's Disease patients. Journal of the Neurological Sciences372, 408-412. https://doi.org/10.1016/j.jns.2016.11.004.
Wang, J. Y., Haider, I., Jamil, M., Fiorilli, V., Saito, Y., Mi, J., Baz, L., Kountche, B.A., Jia, K, P., Guo, X., Balakrishna, A., & Al-Babili, S. (2019). The apocarotenoid metabolite zaxinone regulates growth and strigolactone biosynthesis in rice. Nature Communications10 (1), 810-819. doi.org/10.1038/s41467-019-08461-1.
Yousefi Javan, I., & Gharari, F. (2017). The structure of the protein and gene expression of PIC2 affecting blooming flowers (Crocus sativus L.). Saffron Agronomy & Technology, 5 (1), 73-90. (In Persian with English abstract). https://doi.org/10.22048/jsat.2017.63141.1200.
Yuan, H., Zhang, J., Nageswaran, D., & Li, L. (2015). Carotenoid metabolism and regulation in horticultural crops. Horticulture Research2, 15036-15047. https://doi.org/10.1038/hortres.2015.36.
Yue, J., Wang, R., Ma, X., Liu, J., Lu, X., Thakar, S. B., An, N., Liu, J., Xia, E & Liu, Y. (2020). Full-length transcriptome sequencing provides insights into the evolution of apocarotenoid biosynthesis in Crocus sativusComputational & Structural Biotechnology Journal18, 774-783. https://doi.org/10.1016/j.csbj.2020.03.022.
Zheng, X., Mi, J., Deng, X., & Al-Babili, S. (2021). LC–MS-based profiling provides new insights into apocarotenoid biosynthesis and modifications in citrus fruits. Journal of Agricultural & Food Chemistry69 (6), 1842-1851. https://doi.org/10.1021/acs.jafc.0c06893.