Welcome to Acta Agronomica Sinica,

Acta Agronomica Sinica

   

Heterologous expression of peanut ω-3 fatty acid desaturase genes in Arabidopsis thaliana and transcriptome analysis‌

XU Jing1, BI Jing-Nan1, YIN Xiang-Zhen1, ZHAO Jian-Xin1,2, ZHAO Xu-Hong1, PAN Li-Juan1, CHEN Na1, JIANG Xiao1, MA Jun-Qing1, YIN Dong-Mei3, CHI Xiao-Yuan1,*   

  1. 1 Shandong Peanut Research Institute, Qingdao 266100, Shandong, China;2 College of Biological Engineering, Qingdao University of Science and Technology, Qingdao 266042, Shandong, China; 3 College of Agriculture, Henan Agricultural University, Zhengzhou 450002, Henan, China
  • Received:2025-01-08 Revised:2025-07-09 Accepted:2025-07-09 Published:2025-07-15
  • Supported by:
    This study was supported by the China Agriculture Research System of MOF and MARA (CARS-13), the Major Scientific and Technological Project in Xinjiang (2022A02008-3), the Innovation Project of SAAS (CXGC2025F19), the Peanut Institute Basic Research Tasks Youth Fund General Tasks (CXGC2025C19), the Taishan Scholars Program (tstp20240523, tsqn202312292), the Natural Science Fund of Shandong Province (ZR2023QC177, ZR2023QC146), the Key R&D Program of Shandong Province, China (2024LZGC035), the Open Project of Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture and Rural Affairs, P. R. China (KF2024007), and the Qingdao Natural Science Foundation Youth Project (25-1-1-38-zyyd-jch).

Abstract: The enzyme omega-3 fatty acid dehydrogenase (ω-3 FAD) catalyzes the conversion of linoleic acid to alpha-linolenic acid, a polyunsaturated fatty acid essential for human growth, development, and immune function. To explore the role of ω-3 FAD in peanut growth and development, four ω-3 fatty acid dehydrogenase genes—AhFAD3a, AhFAD3c, AhFAD7a, and AhFAD7d—were cloned from peanut and co-transformed into Arabidopsis thaliana in this study. The results showed that the fatty acid profiles of seeds from transgenic overexpression lines differed significantly from those of the wild type, with total fatty acid content increased by 4.50% to 9.00%. Subcellular localization analysis revealed that AhFAD3a and AhFAD3c were localized to the endoplasmic reticulum, while AhFAD7a and AhFAD7d were targeted to the chloroplast. GO annotation and KEGG pathway enrichment analysis indicated that the differentially expressed genes were significantly enriched in various biological processes, including lipid, amino acid, and carbohydrate metabolism. Notably, 11 upregulated genes were involved in the α-linolenic acid metabolic pathway, and a total of 15 differentially expressed genes associated with fatty acid metabolism were identified. These findings provide a theoretical foundation for understanding the molecular mechanisms of fatty acid metabolism in peanut and offer new insights for the genetic improvement of peanut quality.

Key words: ω-3 fatty acid dehydrogenase, Arabidopsis thaliana, co-expression of multiple gene, transcriptomics, fatty acid metabolism

[1] 万书波. 花生产业形势与对策. 山东农业科学, 2014, 46(10): 128–132.
Wan S B. Situation and developing strategies of peanut industry. Shandong Agric Sci, 2014, 46(10): 128–132 (in Chinese with English abstract).

[2] 赵健鑫, 殷祥贞, 姜骁, 陈娜, 赵旭红, 王明清, 宋虎成, 钟文, 梁成伟, 迟晓元. 不同花生品种烤仁加工特性分析. 花生学报, 2024, 53(2): 91–97.
Zhao J X, Yin X Z, Jiang X, Chen N, Zhao X H, Wang M Q, Song H C, Zhong Wen, Liang C W, Chi X Y. Processing characteristics of roasted kernel of different peanut varieties. J Peanut Sci, 2024, 53(2): 91–97 (in Chinese with English abstract).

[3] 吴列洪, 李付振, 吴学龙, 宋度林, 王美兴. 花生ω-3△15-脂肪酸脱氧酶基因AhFAD3A的克隆及其表达. 中国油料作物学报, 2015, 37: 41–47.
Wu L H, Li F Z, Wu X L, Song D L, Wang M X. Cloning and expression characteristics of one ω-3 cis15fatty acid dehydrogenase gene Ah FAD3A in Arachis hypogaea L. Chin J Oil Crop Sci, 2015, 37: 41–47 (in Chinese with English abstract).

[4] 阮建. 花生ω-3 FAD对提高多不饱和脂肪酸含量的研究. 山东大学硕士毕业论文, 山东济南, 2018.
Ruan J. Functional Research of Peanut ω-3 FAD on Increasing the Content of Polyunsaturated Fatty Acids. MS Thesis of Shandong University, Jinan, Shandong, China, 2018 (in Chinese with English abstract).

[5] Collados R, Andreu V, Picorel R, Alfonso M. A light-sensitive mechanism differently regulates transcription and transcript stability of ω3 fatty-acid desaturases (FAD3, FAD7 and FAD8) in soybean photosynthetic cell suspensions. FEBS Lett, 2006, 580: 4934–4940.

[6] Yang Q Y, Fan C C, Guo Z H, Qin J, Wu J Z, Li Q Y, Fu T D, Zhou Y M. Identification of FAD2 and FAD3 genes in Brassica napus genome and development of allele-specific markers for high oleic and low linolenic acid contents. Theor Appl Genet, 2012, 125: 715–729.

[7] 高继国, 惠识瑶, 耿丽丽, 张蕊, 孙长坡, 张杰. 花生AhFAD8基因的克隆、表达及活性分析. 东北农业大学学报, 2013, 44(10): 82–87.
Gao J G, Hui S Y, Geng L L, Zhang R, Sun C P, Zhang J. Cloning, expression and activity assay of AhFAD8 gene from Arachis hypogaea L. J Northeast Agric Univ, 2013, 44(10): 82–87 (in Chinese with English abstract).

[8] Tovuu A, Zulfugarov I S, Wu G X, Kang I S, Kim C, Moon B Y, An G, Lee C H. Rice mutants deficient in ω-3 fatty acid desaturase (FAD8) fail to acclimate to cold temperatures. Plant Physiol Biochem, 2016, 109: 525–535.

[9] Kodama H, Hamada T, Horiguchi G, Nishimura M, Iba K. Genetic enhancement of cold tolerance by expression of a gene for chloroplast [omega]-3 fatty acid desaturase in transgenic tobacco. Plant Physiol, 1994, 105: 601–605.

[10] Domínguez T, Hernández M L, Pennycooke J C, Jiménez P, Martínez-Rivas J M, Sanz C, Stockinger E J, Sánchez-Serrano J J, Sanmartín M. Increasing omega-3 desaturase expression in tomato results in altered aroma profile and enhanced resistance to cold stress. Plant Physiol, 2010, 153: 655–665.

[11] Chi X Y, Zhang Z M, Chen N, Zhang X W, Wang M, Chen M N, Wang T, Pan L J, Chen J, Yang Z, et al. Isolation and functional analysis of fatty acid desaturase genes from peanut (Arachis hypogaea L.). PLoS One, 2017, 12: e0189759.

[12] 东金玉, 万勇善, 刘风珍. 花生Δ9-硬脂酰-ACP脱氢酶基因(SAD)的序列分析. 作物学报, 2012, 38: 1167–1177.

Dong J Y, Wan Y S, Liu F Z. Sequence analysis of Δ9-Stearoyl-ACP desaturase gene (SAD) in peanut. Acta Agron Sin, 2012, 38: 1167–1177 (in Chinese with English abstract).

[13] Chi X Y, Yang Q L, Pan L J, Chen M N, He Y N, Yang Z, Yu S L. Isolation and characterization of fatty acid desaturase genes from peanut (Arachis hypogaea L.). Plant Cell Rep, 2011, 30: 1393–1404.

[14] Wang Y, Zhang X G, Zhao Y L, Prakash C S, He G H, Yin D M. Insights into the novel members of the FAD2 gene family involved in high-oleate fluxes in peanut. Genome, 2015, 58: 375–383.

[15] Clough S J, Bent A F. Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J, 1998, 16: 735–743.

[16] 陈四龙. 花生油脂合成相关基因的鉴定与功能研究. 中国农业科学院博士学位论文, 北京, 2012.
Chen S L. Idenitfication and Functional Analysis of Lipid Biosynthesis Related Genes in Peanut (Arachis hypogaea L.). PhD Dissertation of Chinese Academy of Agricultural Sciences, Beijing, China, 2012 (in Chinese with English abstract).

[17] Dash S, Cannon E, Kalberer S, Farmer A, Cannon S. Peanuts: Genetics, Processing, and Utilization. New York: Elsevier Inc Academic Press and AOCS Press, 2016. pp 241–252.

[18] Pattee H, Johnss E B, Singleton J A, Sanders T H. Composition changes of peanut fruit parts during maturation. Peanut Sci, 1974, 1: 57–64.

[19] Saini R K, Keum Y S. Omega-3 and omega-6 polyunsaturated fatty acids: Dietary sources, metabolism, and significance: a review. Life Sci, 2018, 203: 255–267.

[20] Alvarenga T I R C, Chen Y Z, Furusho-Garcia I F, Perez J R O, Hopkins D L. Manipulation of omega-3 PUFAs in lamb: phenotypic and genotypic views. Compr Rev Food Sci Food Saf, 2015, 14: 189–204.

[21] Hernández M L, Sicardo M D, Martínez-Rivas J M. Differential contribution of endoplasmic reticulum and chloroplast ω-3 fatty acid desaturase genes to the linolenic acid content of olive (Olea europaea) fruit. Plant Cell Physiol, 2016, 57: 138–151.

[22] Browse J, McConn M, Jr D J, Miquel M. Mutants of Arabidopsis deficient in the synthesis of alpha-linolenate. Biochemical and genetic characterization of the endoplasmic reticulum linoleoyl desaturase. J Biol Chem, 1993, 268: 16345–16351.

[23] 刘训言, 孟庆伟, 李滨. 植物ω-3脂肪酸去饱和酶的研究进展. 细胞生物学杂志, 2004, 26: 34–38.
Liu X Y, Meng Q W, Li B. Advances in ω-3 fatty acid desaturases of plants. Chin J Cell Biol, 2004, 26: 34–38 (in Chinese with English abstract).

[24] Guan L L, Wu W, Hu B, Li D, Chen J W, Hou K, Wang L. Devolopmental and growth temperature regulation of omega-3 fatty acid desaturase genes in safflower (Carthamus tinctorius L.). Genet Mol Res, 2014, 13: 6623–6637.

[25] Bhunia R K, Chakraborty A, Kaur R, Gayatri T, Bhattacharyya J, Basu A, Maiti M K, Sen S K. Seed-specific increased expression of 2S albumin promoter of sesame qualifies it as a useful genetic tool for fatty acid metabolic engineering and related transgenic intervention in sesame and other oil seed crops. Plant Mol Biol, 2014, 86: 351–365.

[26] Wang J J, Liu Z J, Liu H, Peng D S, Zhang J P, Chen M X. Linum usitatissimum FAD2A and FAD3A enhance seed polyunsaturated fatty acid accumulation and seedling cold tolerance in Arabidopsis thaliana. Plant Sci, 2021, 311: 111014.

[27] Liu G, Wu Z H, Shang X H, Peng Y, Gao L Q. Overexpression of PvFAD3 gene from Plukenetia volubilis promotes the biosynthesis of α-linolenic acid in transgenic tobacco seeds. Genes, 2022, 13: 450.

[28] Yeom W W, Kim H J, Lee K R, Cho H S, Kim J Y, Jung H W, Oh S W, Jun S E, Kim H U, Chung Y S. Increased production of α-linolenic acid in soybean seeds by overexpression of Lesquerella FAD3-1. Front Plant Sci, 2020, 10: 1812.

[29] Park M E, Choi H A, Kim H U. Physaria fendleri FAD3-1 overexpression increases ɑ-linolenic acid content in Camelina sativa seeds. Sci Rep, 2023, 13: 7143.

[30] Lee K R, Kim E H, Jeon I, Lee Y, Chen G Q, Kim H U. Lesquerella FAD3-1 gene is responsible for the biosynthesis of trienoic acid and dienoic hydroxy fatty acids in seed oil. Ind Crops Prod, 2019, 134: 257–264.

[31] Li X N, Xue L L, Liu H, Qu P Y, Zhao H H, Luo D D, Wang X B, Huang B Y, Zhang M N, Li C Y, et al. AhFAD3-A01 enhances α-linolenic acid content in Arabidopsis and peanut. Gene, 2025, 949: 149336.

[32] Wakita Y, Otani M, Hamada T, Mori M, Iba K, Shimada T. A tobacco microsomal ω-3 fatty acid desaturase gene increases the linolenic acid content in transgenic sweet potato (Ipomoea batatas). Plant Cell Rep, 2001, 20: 244–249.

[33] 张嘉锡. 油茶亚麻酸合成关键基因CoFAD7的功能分析和优异等位变异挖掘. 中南林业科技大学硕士毕业论文, 湖南长沙, 2024.
Zhang J X. Functional Analysis of CoFAD7, A Key Gene for the Linolenic Acid Synthesis in Camellia Oleifera , and Mining of Excellent Allelic Variation. MS Thesis of Central South University of Forestry & Technology, Changsha, Hunan, China, 2024 (in Chinese with English abstract).

[34] Andreu V, Collados R, Testillano P S, Risueño M D, Picorel R, Alfonso M. In situ molecular identification of the plastid omega3 fatty acid desaturase FAD7 from soybean: evidence of thylakoid membrane localization. Plant Physiol, 2007, 145: 1336–1344.

[35] Andreu V, Lagunas B, Collados R, Picorel R, Alfonso M. The GmFAD7 gene family from soybean: identification of novel genes and tissue-specific conformations of the FAD7 enzyme involved in desaturase activity. J Exp Bot, 2010, 61: 3371–3384.

[36] Peng Z Y, Ruan J, Tian H Y, Shan L, Meng J J, Guo F, Zhang Z M, Ding H, Wan S B, Li X G. The family of peanut fatty acid desaturase genes and a functional analysis of four ω-3 AhFAD3 members. Plant Mol Biol Report, 2020, 38: 209–221.

[37] Zou J, Abrams G D, Barton D L, Taylor D C, Pomeroy M K, Abrams S R. Induction of lipid and oleosin biosynthesis by (+)-abscisic acid and its metabolites in microspore-derived embryos of Brassica napus L.cv Reston (biological responses in the presence of 8 [prime]-hydroxyabscisic acid). Plant Physiol, 1995, 108: 563–571.

[38] Yamamoto K T. Further characterization of auxin-regulated mRNAs in hypocotyl sections of mung bean [Vigna radiata (L.) Wilczek]: sequence homology to genes for fatty-acid desaturases and atypical late-embryogenesis-abundant protein, and the mode of expression of the mRNAs. Planta, 1994, 192: 359–364.

[39] Berberich T, Harada M, Sugawara K, Kodama H, Iba K, Kusano T. Two maize genes encoding omega-3 fatty acid desaturase and their differential expression to temperature. Plant Mol Biol, 1998, 36: 297–306.

[40] Nishiuchi T, Hamada T, Kodama H, Iba K. Wounding changes the spatial expression pattern of the Arabidopsis plastid omega-3 fatty acid desaturase gene (FAD7) through different signal transduction pathways. Plant Cell, 1997, 9: 1701–1712.

[41] Kirsch C, Takamiya-Wik M, Reinold S, Hahlbrock K, Somssich I E. Rapid, transient, and highly localized induction of plastidial omega-3 fatty acid desaturase mRNA at fungal infection sites in Petroselinum crispum. Proc Natl Acad Sci USA, 1997, 94: 2079–2084.

[42] Hajiahmadi Z, Abedi A, Wei H, Sun W B, Ruan H H, Zhuge Q, Movahedi A. Identification, evolution, expression, and docking studies of fatty acid desaturase genes in wheat (Triticum aestivum L.). BMC Genomics, 2020, 21: 778.

[43] Xue Y F, Chen B J, Win A N, Fu C, Lian J P, Liu X, Wang R, Zhang X C, Chai Y R. Omega-3 fatty acid desaturase gene family from two ω-3 sources, Salvia hispanica and Perilla frutescens: Cloning, characterization and expression. PLoS One, 2018, 13: e0191432.

[44] Zoong Lwe Z S, Welti R, Anco D, Naveed S, Rustgi S, Narayanan S. Heat stress elicits remodeling in the anther lipidome of peanut. Sci Rep, 2020, 10: 22163.

[45] 刘婷婷. 玉米苗期干旱复水过程中叶片膜脂响应与干旱适应能力的关系. 中国科学院大学硕士毕业论文, 北京, 2018.
Liu T T. The Relationship between Membrane Lipids Alteration and Drought Adaptation in Leaves of Maize Seedlings. MS Thesis of University of Chinese Academy of Sciences, Beijing, China, 2018 (in Chinese with English abstract).

[46] Zaborowska Z, Starzycki M, Femiak I, Swiderski M, Legocki A B. Yellow lupine gene encoding stearoyl-ACP desaturase--organization, expression and potential application. Acta Biochim Pol, 2002, 49: 29–42.

[47] Nagano M, Kakuta C, Fukao Y, Fujiwara M, Uchimiya H, Kawai-Yamada M. Arabidopsis Bax inhibitor-1 interacts with enzymes related to very-long-chain fatty acid synthesis. J Plant Res, 2019, 132: 131–143.

[1] LI Yan, FANG Yu-Hui, WANG Yong-Xia, PENG Chao-Jun, HUA Xia, QI Xue-Li, HU Lin, XU Wei-Gang. Transcriptomics profile of transgenic OsPHR2 wheat under different phosphorus stress [J]. Acta Agronomica Sinica, 2024, 50(2): 340-353.
[2] DING Hong-Yan, FENG Xiao-Xi, WANG Bai-Yu, ZHANG Ji-Sen. Evolution and relative expression pattern of LRRII-RLK gene family in sugarcane Saccharum spontaneum [J]. Acta Agronomica Sinica, 2023, 49(7): 1769-1784.
[3] WANG Hui-Wei, ZHANG Xiang-Ge, LI Chun-Xin, XU Xin-Ran, HU Hai-Yan, ZHU Ya-Jing, WANG Yan, ZHANG Xin-You. Evaluation of salt tolerance in Cyperus esculentus and transcriptomic analysis of seedling roots under salt stress [J]. Acta Agronomica Sinica, 2023, 49(7): 1882-1894.
[4] ZENG Jian, XU Xian-Chao, XU Yu-Fei, WANG Xiu-Cheng, YU Hai-Yan, FENG Bei-Bei, XING Guang-Nan. Utilization of dynamic transcriptomics analysis for candidate gene mining of 100-seed weight in soybean [J]. Acta Agronomica Sinica, 2021, 47(11): 2121-2133.
[5] Jing DONG,Xiao-Ping LU,Kun-Ming ZHANG,Chun-Lei XUE,Rui-Xia ZHANG. Analysis of SNP and Allele-specific Expression in Transcriptome of Sorghum bicolor × Sorghum sudanense and Their Parents [J]. Acta Agronomica Sinica, 2018, 44(12): 1809-1817.
[6] LIU Ling-Yun,LIU Hao,ZHAO Jing,WANG Yan-Xia,WANG Peng-Tao*. Map-based Cloning and Functional Analysis of Low Chlorophyll Fluorescence Gene LCF3 in Arabidopsis thaliana [J]. Acta Agron Sin, 2016, 42(05): 690-695.
[7] ZHAO Qing-Ping,ZHAO Xiang,MU Shi-Chao,XIAO Hui-Li,ZHANG Xiao. Functional Analysis and Mapping of Gene P2SA2 Involved in Hypocotyl Phototropism of Arabidopsis thaliana [J]. Acta Agron Sin, 2015, 41(04): 585-592.
[8] LIU Jiang,SUN Quan-Xi,LI Xin-Zheng,QI Bao-Xiu. Functional Characterization of Isochrysis galbana Δ5 Desaturase Gene IgD5 in Arabidopsis thaliana [J]. Acta Agron Sin, 2013, 39(05): 928-934.
[9] ZHU Yong-Xing;WANG Lei;ZHANG Lan;ZHANG Wei;FAN Yun-Liu. Isolation and Characterization of Homogentisate Phytyltransferase (HPT) Gene Promoter from Arabidopsis thaliana [J]. Acta Agron Sin, 2007, 33(04): 554-559.
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed   
No Suggested Reading articles found!