Abstract
The precise biological function and activity of the deoxylulose-5-phosphate reductoisomerase (DXR) gene and its promoter in Osmanthus fragrans var. semperflorens remain unclear, even though OfDXR is known as the crucial enzyme involved in plant terpenoid synthesis. This study aimed to shed light on the role and activity of the OfDXR gene and its promoter in O. fragrans var. semperflorens by employing RACE-PCR and Hi-TAIL-PCR techniques for the cloning of the gene and promoter sequence from the petal tissue. Subsequently, genetic transformation and histochemical staining methods were utilized to analyze their function and activity. The OfDXR gene exhibited a DNA sequence length of 5241 bp, encompassing 12 exons and 11 introns. The corresponding cDNA sequence of the OfDXR gene was 1629 bp, encoding 474 amino acid residues. Expression analysis revealed that the OfDXR gene was predominantly active in the petals during the early full blooming stage. Overexpression of the OfDXR gene in Arabidopsis plants at the primary or full blooming stage led to an augmentation in the total terpenoid content. Furthermore, the promoter sequence of the OfDXR gene spanned a length of 1174 bp and contained conserved regulatory/response elements, demonstrating functional activity. These findings indicate that the OfDXR gene plays a pivotal role in terpenoid synthesis, while its promoter exhibits robust activity.
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References
Cai X, Mai RZ, Zou JJ, Zhang HY, Zeng XL, Zheng RR, Wang CY (2014) Analysis of aroma-active compounds in three sweet osmanthus (Osmanthus fragrans) cultivars by GC-olfactometry and GC-MS. J Zhejiang Univ-Sci B 15:638–648. https://doi.org/10.1631/jzus.B1400058
CarreteroPaulet L, Ahumada I, Cunillera N, RodriguezConcepcion M, Ferrer A, Boronat A, Campos N (2002) Expression and molecular analysis of the arabidopsis DXR gene encoding 1-deoxy-d-xylulose 5-phosphate reductoisomerase, the first committed enzyme of the 2-C-methyl-D-erythritol 4-phosphate pathway. Plant Physiol 129:1581–1591. https://doi.org/10.1104/pp.003798
Chen H, Zeng X, Yang J, Cai X, Shi Y, Zheng R, Wang Z, Liu J, Yi X, Xiao S, Fu Q, Zou J, Wang C (2021) Whole-genome resequencing of Osmanthus fragrans provides insights into flower color evolution. Hortic Res 8. https://doi.org/10.1038/s41438-021-00531-0
Deng C, Song G, Hu Y (2004) Application of HS-SPME and GC-MS to characterization of volatile compounds emitted from Osmanthus flowers. Ann Chim 94:921–927. https://doi.org/10.1002/adic.200490114
Devi K, Dehury B, Phukon M, Modi MK, Sen P (2015) Novel insights into structure-function mechanism and tissue-specific expression profiling of full-length dxr gene from Cymbopogon winterianus. Febs Open Bio 5:325–334. https://doi.org/10.1016/j.fob.2015.04.005
Gong Y, Liao Z, Chen M, Zuo K, Guo L, Tan Q, Huang Z, Kai G, Sun X, Tan F, Tang K (2005) Molecular cloning and characterization of a 1-deoxy-D-xylulose 5-phosphate reductoisomerase gene from Ginkgo biloba. DNA Sequence 16:111–120. https://doi.org/10.1080/10425170500058869
Han YJ, Wang HY, Wang XD, Li K, Dong MF, Li Y, Zhu Q, Shang FD (2019) Mechanism of floral scent production in Osmanthus fragrans and the production and regulation of its key floral constituents, β-ionone and linalool. Hortic Res 6. https://doi.org/10.1038/s41438-019-0189-4
Hans J, Hause B, Strack D, Walter MH (2004) Cloning, Characterization, and Immunolocalization of a mycorrhiza-inducible 1-deoxy-D-xylulose 5-phosphate reductoisomerase in arbuscule-containing cells of maize. Plant Physiol 134:614–624. https://doi.org/10.1104/pp.103.032342
Hasunuma T, Takeno S, Hayashi S, Sendai M, Bamba T, Yoshimura S, Tomizawa K, Fukusaki E, Miyake C (2008) Overexpression of 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene in chloroplast contributes to increment of isoprenoid production. J Biosci Bioeng 105:518–526. https://doi.org/10.1263/jbb.105.518
Hu F, Liao X, Chen Z (2018) Determination of three phenylethanoid glycosides in Osmanthus fragrans fruits by high-performance liquid chromatography with fluorescence detection. J Sep Sci 41:3995–4000. https://doi.org/10.1002/jssc.201800610
Lichtenthaler HK (1999) The 1-deoxy-D-xylulose-5-phosphate pathway of isoprenoid biosynthesis in plants. Annu Rev Plant Physiol Plant Mol Biol 50:47–65. https://doi.org/10.1146/annurev.arplant.50.1.47
Liu J, Xu YH, Liang L, Wei JH (2015) Molecular cloning, characterization and expression analysis of the gene encoding 1-deoxy-D-xylulose 5-phosphate reductoisomerase from Aquilaria sinensis (Lour.) Gilg. J Gen 94:239–249. https://doi.org/10.1007/s12041-015-0521-1
Liu YG, Chen Y (2007) High-efficiency thermal asymmetric interlaced PCR for amplification of unknown flanking sequences. Biotechniques 43:649–650. https://doi.org/10.2144/000112601
Mahmoud SS, Croteau RB (2001) Metabolic engineering of essential oil yield and composition in mint by altering expression of deoxyxylulose phosphate reductoisomerase and menthofuran synthase. Proc Natl Acad Sci U S A 98:8915–8920. https://doi.org/10.1073/pnas.141237298
Wang LM, Li MT, Jin WW, Li S, Zhang SQ, Yu LJ (2009) Variations in the components of Osmanthus fragrans Lour. essential oil at different stages of flowering. Food Chem 114:233–236. https://doi.org/10.1016/j.foodchem.2008.09.044
Wu SJ, Shi M, Wu JY (2009) Cloning and characterization of the 1-deoxy-D-xylulose 5-phosphate reductoisomerase gene for diterpenoid tanshinone biosynthesis in Salvia miltiorrhiza (Chinese sage) hairy roots. Biotechnol Appl Biochem 52:89–95. https://doi.org/10.1042/BA20080004
Xin HP, Wu BH, Zhang HH, Wang CY, Li JT, Yang B, Li SH (2013) Characterization of volatile compounds in flowers from four groups of sweet osmanthus (Osmanthus fragrans) cultivars. Can J Plant Sci 93:923–931. https://doi.org/10.4141/Cjps2012-333
Xing S, Miao J, Li S, Qin G, Tang S, Li H, Gu H, Qu LJ (2010) Disruption of the 1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXR) gene results in albino, dwarf and defects in trichome initiation and stomata closure in Arabidopsis. Cell Res 20:688–700. https://doi.org/10.1038/cr.2010.54
Xu C, Li H, Yang X, Gu C, Mu H, Yue Y, Wang L (2016) Cloning and expression analysis of MEP pathway enzyme-encoding genes in Osmanthus fragrans. Genes 7. https://doi.org/10.3390/genes7100078
Xun HW, Zhang X, Yu JM, Pang JS, Wang SC, Liu B, Dong YS, Jiang LL, Guo DQ (2021) Analysis of expression characteristics of soybean leaf and root tissue-specific promoters in Arabidopsis and soybean. Transgenic Res 30:799–810. https://doi.org/10.1007/s11248-021-00266-7
Yao H, Gong Y, Zuo K, Ling H, Qiu C, Zhang F, Wang Y, Pi Y, Liu X, Sun X, Tang K (2008) Molecular cloning, expression profiling and functional analysis of a DXR gene encoding 1-deoxy-D-xylulose 5-phosphate reductoisomerase from Camptotheca acuminata. J Plant Physiol 165:203–213. https://doi.org/10.1016/j.jplph.2006.12.001
Zhang C, Liu H, Hu S, Zong Y, Xia H, Li H (2022) Transcriptomic profiling of the floral fragrance biosynthesis pathway of Liriodendron and functional characterization of the LtuDXR gene. Plant Sci 314:111124. https://doi.org/10.1016/j.plantsci.2021.111124
Zhang C, Wang YG, Fu JX, Bao ZY, Zhao HB (2016) Transcriptomic analysis and carotenogenic gene expression related to petal coloration in Osmanthus fragrans 'Yanhong Gui'. Trees - Struct Funct 30:1207–1223. https://doi.org/10.1007/s00468-016-1359-8
Zhang Y, Yan H, Li Y, Xiong Y, Niu M, Zhang X, TeixeiraSilva JA, Ma G (2021) Molecular cloning and functional analysis of 1-deoxy-D-xylulose 5-phosphate reductoisomerase from Santalum album. Genes 12. https://doi.org/10.3390/genes12050626
Zheng H, Jing L, Yao N, Yang QS, Peng HS, Shen Y, Huang LQ (2016) Cloning and expression analysis of 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene (CcDXR1) in Cinnamomum camphora (L.) Presl. Yao Xue Xue Bao 51:1494–1501
Funding
This research was financially supported by the National Key Research and Development Program of China (No.2017YFD0601201), Key Research and Development Project of Hunan Province (No.2020NK2018), Natural Science Fund of Hunan Province (No.2018JJ3874), and Key Project of Hunan Education Department (No.19B590).
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Z.W. and L.C. conceived the idea and designed the experiment. Z.W., T.H.F., and H.H. conducted the study. Z.W., T.H.F., and A.A.A.S. wrote the manuscript. R.S. gave suggestions to the manuscript. A.F., J.W., and M.S.Y. reviewed and contributed to the discussion. J.W. and L.C. provided the resources and funding.
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Wang, Z., Farooq, T.H., He, H. et al. Cloning and functional analysis of the DXR gene and promoter region in Osmanthus fragrans var. semperflorens. Funct Integr Genomics 23, 277 (2023). https://doi.org/10.1007/s10142-023-01214-w
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DOI: https://doi.org/10.1007/s10142-023-01214-w