Functional Analysis of the PpMYB113 Gene in Anthocyanin Synthesis and Accumulation in Peach (Prunus persica)
ZHOU Shu-Ting1,2, LI Mao-Fu1,3,4, WANG Hua1,3,4, SUN Pei1,3,4, KANG Yan-Hui1,3,4, SUN Xiang-Yi1,3,4, WU Jun-Kai2, ZHANG Li-Bin2,*, JIN Wan-Mei1,3,4,*
1 Institute of Forestry and Pomology, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100093, China; 2 College of Horticultural Science and Technology, Hebei Normal University of Science and Technology, Qinhuangdao 066600; 3 Key Laboratory of Biology and Genetic Improvement of Horticultural Crops (North China), Ministry of Agriculture and Rural Affairs, Beijing 100093, China; 4 Beijing Engineering Research Center for Deciduous Fruit Trees, Beijing 100093, China
Abstract MYB transcription factors play a crucial role in the accumulation of anthocyanins in peach (Prunus persica) fruit. A deeper understanding of MYB transcription factors is beneficial for elucidating the molecular regulatory mechanism of secondary metabolic networks in peach, providing a theoretical basis for genetic improvement, and developing new varieties rich in anthocyanins. In this study, in order to investigate the role of PpMYB113 in the accumulation of anthocyanins in peach flowers and fruits, the PpMYB113 gene (GenBank No. XM_007216468.2) was cloned using red flesh peach as the experimental material. The bioinformatics and expression pattern analysis was performed, and the function of PpMYB113 gene was studied through transient transformation in tobacco (Nicotiana benthamiana) and peach flesh. The results showed that the open reading frame of PpMYB113 gene was 720 bp, encoding 239 amino acids. The phylogenetic tree analysis indicated that the PpMYB113 protein sequence had the closest phylogenetic relationship with the MYB transcription factors of the S6 subfamily in Arabidopsis thaliana. The expression of PpMYB113 gene was analyzed in petals, branches, leaves, bark, fruit skin, and fruit flesh of peach by using semi-quantitative RT-PCR and RT-qPCR. The results indicated that PpMYB113 gene was highly expressed in petals, fruit skin, and fruit flesh, while it was not expressed in branches, leaves, and bark. At the same time, PpMYB113 was overexpressed in tobacco leaves and white fruit flesh using the transient transformation system. The overexpressed tobacco leaves and white fruit flesh showed red color phenotype and the anthocyanin contants were significant increase. In the overexpressed tobacco leaves and white fruit flesh areas, the expression of PpMYB113 was significantly higher than that of the control, and the expression of ANS (anthocyanidin synthase) and UFGT (UDP glucose: flavonoid-3-O-glucosyltransferase) genes were significantly up-regulated. Based on these findings, it was concluded that PpMYB113 plays a positive promoting role in the accumulation of anthocyanins in peach. These results provide a foundation for further investigation into the molecular regulatory mechanism of MYB transcription factors in the accumulation of anthocyanin in red flesh peach.
ZHOU Shu-Ting,LI Mao-Fu,WANG Hua, et al. Functional Analysis of the PpMYB113 Gene in Anthocyanin Synthesis and Accumulation in Peach (Prunus persica)[J]. 农业生物技术学报, 2025, 33(3): 536-546.
[1] 陈静. 2023. SmMYB113调控茄子花脱落的分子机制研究[D]. 硕士学术论文, 山东农业大学, 导师: 杨凤娟, pp. 24-25. (Chen J.2023. Study of the molecular mechanism of SmMYB113 regulate flower abscission in eggplant[D]. Thesis for M.S., Shandong Agricultural University, Supervisor: Yang F J, pp. 24-25.) [2] 李茂福, 杨媛, 王华等. 2022. 月季中R2R3-MYB基因RhMYB113c调控花青苷苷合成[J]. 园艺学报, 49(09): 1957-1966. (Li M F, Yang Y, Wang Het al.2022. The R2R3-MYB gene RhMYB113c regulates anthocyanin synthesis in Chinese rose[J]. Horticultural Plant Journal, 49(09): 1957-1966.) [3] 鲁亚楠. 2023. 文冠果XsMYB113调控花色渐变的分子机制[D]. 博士学术论文, 东北林业大学, 导师: 郑志民. pp. 81-87. (Lu Y N.2023. Molecular mechanism of XsMYB113 regulating yellowhorn flower color change[D]. Thesis for D.S., Northeast Forestry University, Supervisor: Zhen Z M, pp. 81-87.) [4] 莫童. 2021. 亚洲棉黄色花瓣基因GaMYB113的克隆及功能分析[D]. 硕士学术论文, 西南大学, 导师: 肖月华, pp. 28-40. (Mo T.2021. Cloning and functional analysis of yellow petal gene GaMYB113 from Asian cotton[D]. Thesis for M.S., Southwest University, Supervisor: Xiao Y H, pp. 28-40.) [5] 宋志忠, 郭绍雷, 马瑞娟, 等. 2015. KT/HAK/KUP家族基因在桃开花期的表达及对钾肥施用的响应[J]. 中国农业科学, 48(06): 1177-1185. (Song Z Z, Guo S L, Ma R Jet al.2015. Analysis of expression of KT/HAK/KUP family genes and their responses to potassium fertilizer application during peach flowering[J]. Scientia Agricultura Sinica, 48(06): 1177-1185.) [6] 王海波, 周泽宇, 杨振锋. 等. 2023. 我国果业高质量发展的制约因素探析[J]. 中国果树, (07): 1-9. (Wang H B, Zhou Z Y, Yang Z F, et al. 2023. Analysis on the restricting factors of high quality development of fruit industry in China[J]. Chinese Fruit Tree, (07): 1-9.) [7] An G, Chen J.2021. Frequent gain- and loss-of-function mutations of the BjMYB113 gene accounted for leaf color variation in Brassica juncea[J]. BMC Plant Biology, 21(1): 301. [8] An J P, Wang X F, Zhang X W, et al.2020. An apple MYB transcription factor regulates cold tolerance and anthocyanin accumulation and undergoes MIEL1-mediated degradation[J]. Plant Biotechnology Journal, 18: 337-353. [9] Araguirang G E, Richter A S.2022. Activation of anthocyanin biosynthesis in high light - what is the initial signal?[J]. New Phytologist, 236: 2037-2043. [10] Broucke E, Dang T T V, Li Y, et al.2023. SnRK1 inhibits anthocyanin biosynthesis through both transcriptional regulation and direct phosphorylation and dissociation of the MYB/bHLH/TTG1 MBW complex[J]. The Plant Journal, 115(5): 1193-1213. [11] Cao K, Ding T, Mao D, et al.2018. Transcriptome analysis reveals novel genes involved in anthocyanin biosynthesis in the flesh of peach[J]. Plant Physiology Biochemistry, 123: 94-102. [12] Chagné D, Kui L W, Espley R V, et al.2013. An ancient duplication of apple MYB transcription factors is responsible for novel red fruit-flesh phenotypes[J]. Plant Physiology, 161(1): 225-239. [13] Dare A P, Schaffer R J, Kui L W, et al.2008. Identification of a cis-regulatory element by transient analysis of co-ordinately regulated genes[J]. Plant Methods, 4: 17. [14] Drogoudi P, Gerasopoulos D, Kafkaletou M, et al.2017. Phenotypic characterization of qualitative parameters and antioxidant contents in peach and nectarine fruit and changes after jam preparation[J]. Journal of the Science of Food and Agriculture, 97: 3374-3383. [15] Falchi R, Vendramin E, Zanon L, et al.2013. Three distinct mutational mechanisms acting on a single gene underpin the origin of yellow flesh in peach[J]. The Plant Journal, 76: 175-187. [16] Feller A, Yuan L, Grotewold E.2017. The bif domain in plant bHLH proteins is an ACT-like domain[J]. Plant Cell, 29: 1800-1802. [17] Feng R C, Zhang C H, Ma R J, et al.2019. Identification and characterization of WD40 superfamily genes in peach[J]. Gene, 710: 291-306. [18] Feng S Q, Wang Y L, Yang S, et al.2010. Anthocyanin biosynthesis in pears is regulated by a R2R3-MYB transcription factor PyMYB10[J]. Planta, 232: 245-255. [19] Hui Z, Kui L W, Wang F R, et al.2019. Activator-type R2R3- MYB genes induce a repressor-type R2R3-MYB gene to balance anthocyanin and proanthocyanidin accumulation[J]. New Phytologist, 221: 1919-1934. [20] 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. [21] Khan I A, Cao K, Guo J, et al.2022a. Identification of key gene networks controlling anthocyanin biosynthesis in peach flower[J]. Plant Science, 316: 111151. [22] Khan I A, Rahman M U, Sakhi S, et al.2022b. PpMYB39 activates PpDFR to modulate anthocyanin biosynthesis during peach fruit maturation[J]. Horticulturae, 8(4), 332. [23] Kobayashi S, Ishimaru M, Ding C K, et al.2001. Comparison of UDP-glucose: Flavonoid 3-O-glucosyltransferase (UFGT) gene sequences between white grapes (Vitis vinifera) and their sports with red skin[J]. Plant Science, 160: 543-550. [24] Koo Y J, Poethig R S.2021. Expression pattern analysis of three R2R3-MYB transcription factors for the production of anthocyanin in different vegetative stages of Arabidopsis leaves[J]. Applied Biological Chemistry, 64: 1-7. [25] Kreynes A E, Yong Z H, Ellis B E.2021. Developmental phenotypes of Arabidopsis plants expressing phosphovariants of AtMYB75[J]. Plant Signal & Behavior, 16(1): e1836454. [26] Kui L W, 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: 50. [27] Liu H, Xiong J S, Jiang Y T, et al.2019. Evolution of the R2R3-MYB gene family in six Rosaceae species and expression in woodland strawberry[J]. Journal of Integrative Agriculture, 18: 2753-2770. [28] Liu X F, Feng C, Zhang M M, et al.2013. The MrWD40-1 gene of Chinese bayberry (Myrica rubra) interacts with MYB and bHLH to enhance anthocyanin accumulation[J]. Plant Molecular Biology Reporter, 31: 1474-1484. [29] Liu Y, Tikunov Y, Schouten R E, et al.2018. Anthocyanin biosynthesis and degradation mechanisms in Solanaceous vegetables: A review[J]. Frontiers Chemistry, 6: 52. [30] Seo M S, Kim J S.2017. Understanding of MYB transcription factors involved in glucosinolate biosynthesis in Brassicaceae[J]. Molecules, 22(9): 1549. [31] Tuan P A, Bai S, Yaegaki H, et al.2015. The crucial role of PpMYB10.1 in anthocyanin accumulation in peach and relationships between its allelic type and skin color phenotype[J]. BMC Plant Biology 15: 280. [32] Uematsu C, Katayama H, Makino I, et al.2014. Peace, a MYB-like transcription factor, regulates petal pigmentation in flowering peach 'Genpei' bearing variegated and fully pigmented flowers[J]. Journal of Experimental Botany, 65: 1081-1094. [33] Wang H, Zhang H, Yang Y, et al.2020. The control of red colour by a family of MYB transcription factors in octoploid strawberry (Fragaria×ananassa) fruits[J]. Plant Biotechnology Journal 18, 1169-1184. [34] Xu W, Dubos C, Lepiniec L.2015. Transcriptional control of flavonoid biosynthesis by MYB-bHLH-WDR complexes[J]. Trends in Plant Science, 20: 176-185. [35] Yan H L, Pei X N, Zhang H, et al.2021. MYB-mediated regulation of anthocyanin biosynthesis[J]. International Journal of Molecular Sciences, 22(6): 3103. [36] Ying H, Shi J, Zhang S, et al.2019. Transcriptomic and metabolomic profiling provide novel insights into fruit development and flesh coloration in Prunus mira Koehne, a special wild peach species[J]. BMC Plant Biology, 19: 463. [37] Yuan Y, Li X, Yao X, et al.2023. Mechanisms underlying the formation of complex color patterns on Nigella orientalis (Ranunculaceae) petals[J]. New Phytologist, 237(6): 2450-2466. [38] Zhao X, Zhang W, Yin X, et al.2015. Phenolic composition and antioxidant properties of different peach [Prunus persica (L.) Batsch] cultivars in China[J]. International Journal of Molecular Sciences, 16: 5762-5778. [39] Zhou H, Kui L W, Wang H L, et al.2015. Molecular genetics of blood-fleshed peach reveals activation of anthocyanin biosynthesis by NAC transcription factors[J]. Plant Journal, 82: 105-121. [40] Zhou L, He Y, Li J, et al.2020. CBFs Function in anthocyanin biosynthesis by interacting with MYB113 in eggplant (Solanum melongena L.)[J]. Plant Cell Physiology, 61(2):416-426. [41] Zhou Y, Zhou H, Kui L W, et al.2014. Transcriptome analysis and transient transformation suggest an ancient duplicated MYB transcription factor as a candidate gene for leaf red coloration in peach[J]. BMC Plant Biology, 14: 388.
