白木香AsABCG1基因的克隆与表达分析

Cloning and expression analysis of AsABCG1 gene in Aquilaria sinensis

摘要

沉香是一种传统的名贵药材与天然香料，因蕴藏丰富的次生代谢产物而极具价值。ABC 转运蛋白是一类广泛分布于生物体的跨膜蛋白，在植物次生代谢产物的运输与积累中起着关键作用。为探究 ABC 转运蛋白在沉香次生代谢物转运过程中的功能，通过分析白木香（Aquilaria sinensis）转录组数据，鉴定出一个编码 ABC 转运蛋白的基因 AsABCG1，该基因编码区全长为 1 986 bp，编码一个由 661 个氨基酸组成的蛋白，AsABCG1 具有典型的植物 ABC 转运蛋白结构特征，包含两个保守结构域 MotifA 和 MotifB。AsABCG1 启动子区含有茉莉酸、生长素、赤霉素、脱落酸等多种激素响应元件及厌氧诱导、机械损伤等胁迫响应元件。实时荧光定量 PCR（RT-qPCR）分析表明，茉莉酸甲酯和机械伤害处理均能够显著诱导白木香茎干中AsABCG1 的表达。本研究为后续阐明 AsABCG1 基因在白木香防御胁迫响应和次生代谢物积累中的功能奠定了基础。

Abstract

Agarwood is a traditional precious medicinal material and a natural aromatic material, which is highly valuable due to its rich secondary metabolites. ABC transporters are a type of transmembrane proteins widely distributed in organisms and play a crucial role in the transport and accumulation of secondary metabolites in plants. To deeply explore the function of ABC transporters in the transport process of secondary metabolites in agarwood, a gene encoding an ABC transporter AsABCG1; was identified by analyzing the transcriptome data of Aquilaria sinensis. The coding sequence (CDS) length of this gene is 1986 bp, encoding a protein consisting of 661 amino acids. AsABCG1 has typical structural characteristics of plant ABC transporters, including two conserved domains, Motif A and Motif B. The promoter region of AsABCG1 contains various hormone response elements such as jasmonic acid, auxin, gibberellin, abscisic acid, as well as stress response elements such as anaerobic induction and mechanical injury. The real-time fluorescence quantitative PCR (RT-qPCR) analysis showed that methyl jasmonic acid and mechanical injury treatments could significantly induce the expression of AsABCG1 in the stems of A. sinensis. All these findings laid the foundation for further elucidating the function of the AsABCG1 gene in the defense response to stress and the accumulation of secondary metabolites in A. sinensis.

关键词

白木香 / ABC 转运蛋白 / AsABCG1 / 表达分析 / 次生代谢物

Key words

Aquilaria sinensis / ABC transporter protein / AsABCG1 / expression analysis / secondary metabolite

参考文献

[1] 李慧婷, 王露露, 王雅丽, 等. 白木香 6 个不同种质所结沉香的 GC-MS 分析[J]. 热带作物学报, 2024, 45(8): 1727-1741. https://doi.org/10.3969/j.issn.1000-2561.2024.08.022

[2] Zhou H, Li X Y, Fang H B, et al. Five new sesquiterpenoids from agarwood of Aquilaria sinensis[J]. Beilstein Journal of Organic Chemistry, 2023, 19: 998-1007. https://doi.org/10.3762/bjoc.19.75

[3] Zhang H, Ma J L, Chang C, et al. Gastroprotective 2-(2-phenylethyl) chromone-sesquiterpene hybrids from the resinous wood of Aquilaria sinensis (Lour.) Gilg[J]. Bioorganic Chemistry, 2023, 133: 106396. https://doi.org/10.1016/j.bioorg.2023.106396

[4] Xia L L, Li W, Wang H, et al. LC-MS guided identification of dimeric 2-(2-phenylethyl) chromones and sesquiterpene-2-(2-phenylethyl) chromone conjugates from agarwood of Aquilaria crassna and their cytotoxicity[J]. Fitoterapia, 2019, 138: 104349. https://doi.org/10.1016/j.fitote.2019.104349

[5] Li C G, Pan L, Han Z Z, et al. Antioxidative 2-(2-phenylethyl) chromones in Chinese eaglewood from Aquilaria sinensis[J]. Journal of Asian Natural Products Research, 2020, 22(7): 639-646. https://doi.org/10.1080/10286020.2019.1607841

[6] Wei S Y, Hu D B, Xia M Y, et al. Sesquiterpenoids and 2-(2-phenylethyl) chromone derivatives from the resinous heartwood of Aquilaria sinensis[J]. Natural Products and Bioprospecting, 2021, 11(5): 545-555. https://doi.org/10.1007/s13659-021-00313-0

[7] Wang Z F, Cao H L. The complete mitochondrial genome sequence ofAquilaria sinensis[J]. Mitochondrial DNA Part B, 2021, 6(2): 381-383. https://doi.org/10.1080/23802359.2020.1869609

[8] Chen Y, Wu K L, Xu J R, et al. Development and application of SSR markers for Aquilaria sinensis on the basis of whole-genome resequencing data[J]. Plants, 2025, 14(9): 1323. https://doi.org/10.3390/plants14091323

[9] 黄兴, 梅文莉, 王昊, 等. 白木香 AsCYP131 基因的克隆及表达分析[J]. 热带作物学报, 2025, 46(5): 1076-1083. https://doi.org/10.3969/j.issn.1000-2561.2025.05.006

[10] Naziz P S, Das R, Sen S. The scent of stress: evidence from the unique fragrance of agarwood[J]. Frontiers in Plant Science, 2019, 10: 840. https://doi.org/10.3389/fpls.2019.00840

[11] Alam A, Locher K P. Structure and mechanism of human ABC transporters[J]. Annual Review of Biophysics, 2023, 52: 275-300. https://doi.org/10.1146/annurev-biophys-111622-091232

[12] Locher K P. Mechanistic diversity in ATP-binding cassette (ABC) transporters[J]. Nature Structural & Molecular Biology, 2016, 23(6): 487-493. https://doi.org/10.1038/nsmb.3216

[13] Dhara A, Raichaudhuri A. ABCG transporter proteins with beneficial activity on plants[J]. Phytochemistry, 2021, 184: 112663. https://doi.org/10.1016/j.phytochem.2021.112663

[14] Lewinson O, Orelle C, Seeger M A. Structures of ABC transporters: handle with care[J]. FEBS Letters, 2020, 594(23): 3799-3814. https://doi.org/10.1002/1873-3468.13966

[15] Yan L, Zhang J H, Chen H Y, et al. Genome-wide analysis of ATP-binding cassette transporter provides insight to genes related to bioactive metabolite transportation in Salvia miltiorrhiza[J]. BMC Genomics, 2021, 22(1): 315. https://doi.org/10.1186/s12864-021-07623-0

[16] Ichino T, Yazaki K. Modes of secretion of plant lipophilic metabolites via ABCG transporter-dependent transport and vesicle-mediated trafficking[J]. Current Opinion in Plant Biology, 2022, 66: 102184. https://doi.org/10.1016/j.pbi.2022.102184

[17] LI R S, ZHU J H, GUO D, et al. Genome-wide identification and expression analysis of terpene synthase gene family in Aquilaria sinensis[J]. Plant Physiology and Biochemistry, 2021, 164: 185-194. https://doi.org/10.1016/j.plaphy.2021.04.028

[18] 梅文莉, 戴好富, 王辉, 等. 一种生产沉香的方法: 中国, 201310150138.5[P]. 2013-04-26. https://d.wanfangdata.com.cn/patent/Ch1QYXRlbnROZXdTb2xyOVMyMDI2MDEyNzE1NTEzORIpWkxfQ04yMDEzMTAxNTAxMzguNV9DTjEwMzIyOTY3N0JfMjAxNDA1MTQaCDgzbGlxemIy.

[19] How S S, Nathan S, Lam S D, et al. ATP-binding cassette (ABC) transporters: structures and roles in bacterial pathogenesis[J]. Journal of Zhejiang University-SCIENCE B, 2025, 26(1): 58-75. https://doi.org/10.1631/jzus.B2300641

[20] Li Y J, Chen J F, Zhi J Y, et al. The ABC transporter SmABCG1 mediates tanshinones export from the peridermic cells of Salvia miltiorrhiza root[J]. Journal of Integrative Plant Biology, 2025, 67(1): 135-149. https://doi.org/10.1111/jipb.13806

[21] Lin G Y, Li P R, Li L W, et al. Discovery of ElABCG39: a key player in ingenol transmembrane efflux identified through genome-wide analysis of ABC transporters in Euphorbia lathyris L.[J]. Plant Cell Reports, 2024, 43(11): 274. https://doi.org/10.1007/s00299-024-03361-1

[22] Dean J V, Willis M, Shaban L. Transport of acylated anthocyanins by the Arabidopsis ATP-binding cassette transporters AtABCC1, AtABCC2, and AtABCC14[J]. Physiologia Plantarum, 2022, 174(5): e13780. https://doi.org/10.1111/ppl.13780

[23] Zhang M, Zhao Y Y, Nan T G, et al. Genome-wide analysis of Citrus medica ABC transporters reveals the regulation of fruit development by CmABCB19 and CmABCC10[J]. Plant Physiology and Biochemistry, 2024, 215: 109027. https://doi.org/10.1016/j.plaphy.2024.109027

[24] 王昊, 丁旭坡, 曾军, 等. 沉香种质资源、品质评价与形成机制研究进展[J]. 中国科学: 生命科学, 2024, 54(10): 1885-1906. https://doi.org/10.1360/SSV-2023-0291

[25] Ding X P, Mei W L, Lin Q, et al. Genome sequence of the agarwood tree Aquilaria sinensis (Lour.) Spreng: the first chromosome-level draft genome in the Thymelaeaceae family[J]. GigaScience, 2020, 9(3): giaa013. https://doi.org/10.1093/gigascience/giaa013

[26] Xu J R, Du R Y, Wang Y, et al. Wound-induced temporal reprogramming of gene expression during agarwood formation in Aquilaria sinensis[J]. Plants, 2023, 12(16): 2901. https://doi.org/10.3390/plants12162901

[27] 张鹏, 徐谙为, 杨光大, 等. 物理伤害诱导白木香应激反应的生理特征[J]. 热带作物学报, 2021, 42(12): 3521-3528. https://doi.org/10.3969/j.issn.1000-2561.2021.12.019

[28] Xu Y H, Sun P W, Tang X L, et al. Genome-wide analysis of WRKY transcription factors in Aquilaria sinensis (Lour.) Gilg[J]. Scientific Reports, 2020, 10(1): 3018. https://doi.org/10.1038/s41598-020-59597-w

基金资助

国家自然科学基金项目(32171747)

海南省自然科学基金项目(323CXTD388)