Effect of FaMYB10 Allele Nucleotide Polymorphism on Anthocyanin Accumulation in Tobacco (Nicotiana benthamiana)
WANG Hua1,2, DONG Jing1,2,3, YANG Yuan1,2,3, LI Mao-Fu1,2, SUN Pei1,2, FAN You-Wei1,2, WANG Gui-Xia1,2,3,*, JIN Wan-Mei1,2,*
1 Beijing Academy of Forestry and Pomology Sciences, Beijing 100093, China; 2 Key Laboratory of Biology and Genetic Improvement of Horticultural Crops (North China), Ministry of Agriculture and Rural Affairs, Beijing 100093, China; 3 Beijing Engineering Research Center for Deciduous Fruit Trees, Beijing 100093, China
Abstract:Anthocyanins are important pigments for strawberry (Fragaria × ananassa) fruit coloring, and FAMYB10 is a key transcription factor for anthocyanin synthesis. To investigate the role of nucleotide polymorphisms of FaMYB10 alleles in anthocyanin accumulation in tobacco (Nicotiana benthamiana), in this study, 3 alleles of FaMYB10 were isolated from the fruit by homologous cloning. FaMYB10a (GenBank No. MW478285) and FaMYB10b (GenBank No. MG456859) were identified from the red strawberry variety 'Sweet Charlie', and FaMYB10c (GenBank No. MG456860) was identified in the fruit of the white variety 'Snow White'. Sequence analysis showed that the open reading frame of FaMYB10a and FaMYB10b was 702 bp, encoding 233 amino acids. There was a single nucleotide polymorphism at 94 bp from start codon ATG, where FaMYB10a was base A, and FaMYB10b was base C. The open reading frame of FaMYB10c was 710 bp, and there was a 8 base insertions at 491 bp starting from ATG, which led to premature termination and the encoding of 179 amino acids. The bases insertion in the open reading frame of FaMYB10c resulted in differences in physicochemical properties and protein structure from the other 2 alleles. When FaMYB10a and FaMYB10b were overexpressed, the tobacco plants showed anthocyanin accumulation and a red phenotype. However, when FaMYB10c was overexpressed, the tobacco plants showed no anthocyanin accumulation. The above results indicated that the nucleotide polymorphisms of FaMYB10a and FaMYB10b did not affect the anthocyanin accumulation, while the nucleotide polymorphisms of FaMYB10c led to functional changes, and ultimately tobacco leaves could not accumulate anthocyanins. The present study provides reference for the functional research of other important genes, and also provides a reference for the study of plant traits and evolution.
王华, 董静, 杨媛, 李茂福, 孙佩, 范又维, 王桂霞, 金万梅. FaMYB10等位基因核苷酸多态性对烟草花青素积累的作用[J]. 农业生物技术学报, 2021, 29(7): 1274-1282.
WANG Hua, DONG Jing, YANG Yuan, LI Mao-Fu, SUN Pei, FAN You-Wei, WANG Gui-Xia, JIN Wan-Mei. Effect of FaMYB10 Allele Nucleotide Polymorphism on Anthocyanin Accumulation in Tobacco (Nicotiana benthamiana). 农业生物技术学报, 2021, 29(7): 1274-1282.
[1] 葛翠莲, 黄春辉, 徐小彪.2012.果实花青素生物合成研究进展[J].园艺学报, 39(9): 1655-1664. (Ge C L, Huang C H, Xu X B.2012.Research on anthocyanins biosynthesis in fruit[J].Acta Horticulturae Sinica, 39(9): 1655-1664.) [2] 刘晓芬, 李方, 殷学仁, 等.2013.花青苷生物合成转录调控研究进展[J].园艺学报, 40(11): 2295-2306. (Liu X F, Li F,Yin X R, et al.2013.Recent advances in the transcriptional regulation of anthocyanin biosynthesis[J].Acta Horticulturae Sinica, 40(11): 2295-2306.) [3] 王华, 李茂福, 杨媛, 等.2015.果实花青素生物合成分子机制研究进展[J].植物生理学报, 51(1): 29-43. (Wang H, Li M F, Yang Y, et al.2015.Recent advances on the molecular mechanisms of anthocyanin synthesis in fruits[J].Plant Physiology Journal, 51(1): 29-43.) [4] 武晶, 黎裕.2019.基于作物种质资源的优异等位基因挖掘:进展与展望[J].植物遗传资源学报, 20(6): 1380-1389. (Wu J, Li Y.2019.Mining superior alleles in crop germplasm resources: Advances and perspectives[J].Journal of Plant Genetic Resources, 20(6): 1380-1389.) [5] Allan A C, Hellens R P, Laing W A.2008.MYB transcription factors that colour our fruit[J].Trends in Plant Science,13(3): 99-102. [6] Ban Y, Honda C, Hatsuyama Y, et al.2007.Isolation and functional analysis of a MYB transcription factor gene that is a key regulator for the development of red coloration in apple skin[J].Plant and Cell Physiology, 48(7): 958-970. [7] Benvenuti S, Pellati F, Melegari M, et al.2004.Polyphenols, anthocyanins, ascorbic acid, and radical scavenging activity of Rubus, Ribes, and Aronia[J].Journal of Food Science, 69(3):164-169. [8] Butelli E, Garcia-Lor A, Licciardello C, et al.2017.Changes in anthocyanin production during domestication of Citrus[J].Plant Physiology,173(4): 2225-2242. [9] Castillejo C, Waurich V, Wagner H, et al.2020.Allelic variation of MYB10 is the major force controlling natural variation in skin and flesh color in strawberry (Fragaria spp.) fruit[J].The Plant Cell, 32(12): 3723-3749. [10] Dubos C, Stracke R, Grotewold E, et al.2010.MYB transcription factors in Arabidopsis[J].Trends in Plant Science, 15(10): 573-581. [11] Edger P P, PoortenT J, VanBuren R, et al.2019.Origin and evolution of the octoploid strawberry genome[J].Nature genetics, 51(3): 541-547. [12] Espley R V, Hellens R P, Putterill J, et al.2007.Red colouration in apple fruit is due to the activity of the MYB transcription factor, MdMYB10[J].The Plant Journal, 49(3): 414-427. [13] Feng C, Wang J, Harris A J, et al.2021.Tracing the diploid ancestry of the cultivated octoploid strawberry[J].Molecular Biology and Evolution, 38(2): 478-485. [14] Gao L, Gonda I, Sun H, et al.2019.The tomato pan-genome uncovers new genes and a rare allele regulating fruit flavor[J].Nature genetics, 51(6): 1044-1051. [15] Hichri I, Heppel S C, Pillet J, et al.2010.The basic helix-loop-helix transcription factor MYC1 is involved in the regulation of the flavonoid biosynthesis pathway in grapevine[J].Molecular Plant, 3(3): 509-523. [16] Hichri I, Barrieu F, Bogs J, et al.2011.Recent advances in the transcriptional regulation of the flavonoid biosynthetic pathway[J].Journal of Experimental Botany, 62(8): 2465-2483. [17] Jaakola L.2013.New insights into the regulation of anthocyanin biosynthesis in fruits[J].Trends in Plant Science, 18(9): 477-483. [18] Jin W, Wang H, Li M, et al.2016.The R2R3 MYB transcription factor PavMYB10.1 involves in anthocyanin biosynthesis and determines fruit skin colour in sweet cherry (Prunus avium L.)[J].Plant Biotechnology Journal, 14(11): 2120-2133. [19] Jiang L, Ma X, Zhao S, et al.2019.The APETALA2-like transcription factor SUPERNUMERARY BRACT controls rice seed shattering and seed size[J].The Plant Cell, 31(1): 17-36. [20] Kobayashi S, Goto-Yamamoto N, Hirochika H.2004.Retrotransposon-induced mutations in grape skin color[J].Science, 304(5673): 982. [21] Koeda S, Nakano R, Sawaki T, et al.2020.Multiple non-pungent Capsicum chinense accessions with a loss of function CaKR1 allele originating from south America[J].The Horticulture Journal.DOI: 10.2503/hortj.UTD-184. [22] Kong J M, Chia LS, Goh N K, et al.2003.Analysis and biological activities of anthocyanins[J].Phytochemistry, 64(5): 923-933. [23] Li Y, Zhu J, Wu H, et al.2020.Precise base editing of non-allelic acetolactate synthase genes confers sulfonylurea herbicide resistance in maize[J].The Crop Journal, 8(3): 449-456. [24] Lin-Wang K, Bolitho K, Grafton K, et al.2010.An R2R3 MYB transcription factor associated with regulation of the anthocyanin biosynthetic pathway in Rosaceae[J].BMC Plant Biology, 10(1): 50. [25] Sainsbury F, Thuenemann E C, Lomonossoff G P.2009.pEAQ: Versatile expression vectors for easy and quick transient expression of heterologous proteins in plants[J].Plant Biotechnology Journal, 7(7): 682-693. [26] Schaart J G, Dubos C, Romero De La Fuente I, et al.2013.Identification and characterization of MYB-bHLH-WD40 regulatory complexes controlling proanthocyanidin biosynthesis in strawberry (Fragaria × ananassa) fruits[J].New Phytologist, 197(2): 454-467. [27] Sheick R, Serra S, Tillman J, et al.2020.Characterization of a novel S-RNase allele and genotyping of new apple cultivars[J].Scientia Horticulturae, 273: 109630. [28] This P, Lacombe T, Cadle-Davidson M, et al.2007.Wine grape (Vitis vinifera L.) color associates with allelic variation in the domestication gene VvmybA1[J].Theoretical and Applied Genetics, 114(4): 723-730. [29] Walker A R, Lee E, Bogs J, et al.2007.White grapes arose through the mutation of two similar and adjacent regulatory genes[J].The Plant Journal, 49(5): 772-785. [30] Wang D, Pang Y, Dong L, et al.2020a.Allelic impacts on pre-harvest sprouting resistance and favorable haplotypes in TaPHS1 of Chinese wheat accessions[J].The Crop Journal, 8: 515-521. [31] Wang H, Zhang H, Yang Y, et al.2020b.The control of red colour by a family of MYB transcription factors in octoploid strawberry (Fragaria × ananassa) fruits[J].Plant Biotechnology Journal, 18(5): 1169-1184. [32] YangY, Cuenca J, Wang N, et al.2020.A key 'foxy' aroma gene is regulated by homology-induced promoter indels in the iconic juice grape 'Concord'[J].Horticulture Research, 7(1): 1-12. [33] Zhang L, Hu J, Han X, et al.2019.A high-quality apple genome assembly reveals the association of a retrotransposon and red fruit colour[J].Nature Communications, 10(1): 1-13. [34] Zhao J, Jiang L, Che G, et al.2019.A functional allele of CsFUL1 regulates fruit length through repressing CsSUP and inhibiting auxin transport in cucumber[J].The Plant Cell, 31(6): 1289-1307.