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| Identification of the bHLH Gene Family in Kiwifruit (Actinidia chinensis) and Its Expression Analysis Under Fruit Storage Conditions |
| ZHANG Shi-Ying, ZHOU Liang, MAO Jia-Xu, WANG Lu-Fan, SHEN Shu-Ling, ZHENG Xiao-Lin, HUAN Chen* |
| College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou 310018, China |
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Abstract Kiwifruit is rich in ascorbic acid (AsA), but the AsA content significantly decreases during postharvest ripening. AsA metabolism is regulated by transcription factors. The basic helix-loop-helix (bHLH) gene family has been proven to be involved in various metabolic pathways. To explore the role of the bHLH gene family in AsA metabolism in kiwifruit, this study conducted bioinformatics identification of the kiwifruit (Actinidia chinensis) bHLH gene family. The physicochemical properties, chromosomal localization, conserved motifs, evolutionary relationships, synonymous codon usage frequency, and protein interaction networks of these members were analyzed to determine gene nomenclature and classify the gene family, predicting their interaction relationships. Finally, by analyzing the expression patterns of bHLH transcription factors (TFs) and AsA metabolic genes during postharvest storage, combined with the prediction of cis-acting elements in the promoters of functional genes, the potential links between bHLH TFs and AsA metabolism were inferred. The results showed that the 144 members were distributed across 29 chromosomes, with most members possessing typical bHLH conserved motifs. The cis-acting elements included 7 major categories: Light response, stress response, MYB binding, hormone response, defense and stress response, endosperm and meristem expression and circadian rhythm. According to the phylogenetic tree analysis, 144 bHLH members were divided into 25 subfamilies, of which 7 members having a relative synonymous codon usage frequency greater than 2.0, indicating a strong codon usage preference in the AcbHLH gene family. Protein interaction network analysis revealed interactions among 60 members. qRT-PCR analysis indicated that AcbHLH072, AcbHLH120, AcbHLH134, and AcbHLH067 were significantly correlated with AsA content. Additionally, 8 AsA metabolism-related genes all contained cis-acting elements for bHLH binding, suggesting that AcbHLH072, AcbHLH120, AcbHLH134, and AcbHLH067 might regulate AsA synthesis and degradation by influencing the expression of related genes such as ascorbate peroxidase 2 (AcAPX2), AcAPX8, L-galactose dehydrogenase 1 (AcGalDH1), galacturonic acid uridylyltransferase 2 (AcGalUR2), and AcGalUR3, thereby modulating the AsA metabolic pathway in the fruit. This study provides insights into the mechanisms by which the kiwifruit bHLH gene family participates in ascorbic acid metabolism.
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Received: 10 March 2025
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Corresponding Authors:
*huanchen@zjgsu.edu.cn
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[1] 安昌, 陆琳, 沈梦千, 等. 2023. 植物bHLH基因家族研究进展及在药用植物中的应用前景[J]. 生物技术通报, 39(10): 1-16.
(An C, Lu L, Shen M Q, et al.2018.Research progress on the plant bHLH gene family and its application prospects in medicinal plants[J]. Biotechnology Bulletin, 39(10): 1-16.)
[2] 陈红霖, 胡亮亮, 王丽侠, 等. 2017. 绿豆bHLH转录因子家族的鉴定与生物信息学分析[J]. 植物遗传资源学报, 18(06): 1159-1167.
(Chen H L, Hu L L, Wang L X, et al.2017. Identification and bioinformatics analysis of the bHLH transcription factor family in mung bean[J]. Journal of Plant Genetic Resources, 18(06): 1159-1167.)
[3] 董慧珍. 2020. 梨bHLH基因家族生物信息学分析及PbbHLH67基因耐盐性鉴定[D]. 硕士学位论文, 南京农业大学, 导师: 黄小三, pp. 39-41.
(Dong H Z.2020. Bioinformaties analysis of pear bHLH family and identification of PbbHLH67 gene salt resistance[D]. Thesis for M.S., Nanjing Agricultural University, Supervisor: Huang X S, pp. 39-41.)
[4] 郭琳琳, 庞荣丽, 王瑞萍, 等. 2022. 猕猴桃营养品质综合评价[J]. 果树学报, 39(10): 1864-1872.
(Guo L L, Pang R L, Wang R P, et al.2022. Comprehensive evaluation of the nutritional quality of kiwifruit[J]. Journal of Fruit Science, 39(10): 1864-1872.)
[5] 梁华俤, 潘林娜, 魏勤, 等. 1993. 猕猴桃果实采后Vc变化规律及其稳定性[J]. 食品科学, 14(5): 3-9.
(Liang H D, Pan L N, Wei Q, et al.1993. The changing rules and stability of Vc of plucked kiwi[J]. Food Science, 14(5): 3-9.)
[6] 刘菁菁, 王舒文, 杨旭锐, 等. 2025. bHLH转录因子在植物花青素合成中的调控作用[J/OL]. 植物遗传资源学报, 1-16.
(Liu Q Q, Wang S W, Yang X R, et al.2025. The regulatory role of bHLH transcription factors in the synthesis of plant anthocyanins[J/OL]. Journal of Plant Genetic Resources, 1-16.)
[7] 刘硕. 2022. 软枣猕猴桃AaMYB16转录因子响应冻胁迫的功能解析[D]. 硕士学位论文, 东北林业大学, 导师: 孟凡娟, pp. 50.
(Liu S.2022. The functional analysis of AaMYB16 transcriptionfactor responses to freezing tolerance in Actinidia arguta[D]. Thesis for M.S., Northeast Forestry University, Supervisor: Meng F J, pp. 50.)
[8] 李正男, 陈磊, 张立培, 等. 2022. 玉米CKX基因家族生物信息学及表达模式分析[J]. 分子植物育种, 20(13): 4195-4205.
(Li Z N, Chen L, Zhang L P, et al.2022. Bioinformatics and expression pattern analysis of the corn CKX gene family[J]. Molecular Plant Breeding, 20(13): 4195-4205.)
[9] 夏明辉, 张珅, 赵云峰, 等. 2023. 外源γ-氨基丁酸延缓采后猕猴桃果实冷害及其与活性氧代谢的关系[J]. 食品科学, 44(19): 198-206.
(Xia M H, Zhang K, Zhao Y F, et al.Exogenous γ-aminobutyric acid delays postharvest kiwifruit cold injury and its relationship with reactive oxygen metabolism[J]. Food Science, 44(19): 198-206.)
[10] 王昕嘉, 李昆志. 2015. 植物bHLH转录因子参与非生物胁迫信号通路研究进展[J]. 生命科学, 27(02): 208-216.
(Wang X J, Li K Z.2015. Research progress on the participation of plant bHLH transcription factors in abiotic stress signaling pathways[J]. Chinese Bulletin of Life Sciences, 27(02): 208-216.)
[11] 杨朝结, 张兰, 陈红, 等. 2025. 苦荞转录因子基因FtbHLH3调控类黄酮生物合成的功能鉴定[J/OL]. 生物技术通报, 1-11.
(Yang C J, Zhang L, Chen H, et al.2025. Functional identification of the transcription factor gene FtbHLH3 in regulating flavonoid biosynthesis in Fagopyrum tataricum[J/OL]. Biotechnology Bulletin, 1-11.)
[12] Chen Y, Shu P, Wang R C, et al.2021. Ethylene response factor AcERF91 affects ascorbate metabolism via regulation of GDP-galactose phosphorylase encoding gene (AcGGP3) in kiwifruit[J]. Plant Science, 313: 111063.
[13] Gao F, Dubos C.2023. The Arabidopsis bHLH transcription factor family[J]. Trends in Plant Science, 29(6): 668-680.
[14] He X, Zhang W, Sabir I A, et al.2023. The spatiotemporal profile of Dendrobium huoshanense and functional identification of bHLH genes under exogenous MeJA using comparative transcriptomics and genomics[J]. Frontiers in Plant Science, 14: 1169386.
[15] Huan C, Du X J, Wang L F, et al.2021.Transcriptome analysis reveals the metabolisms of starch degradation and ethanol fermentation involved in alcoholic off-flavour development in kiwifruit during ambient storage[J]. Postharvest Biology and Technology, 180: 111621.
[16] Huang Q M, Yan Y L, Zhang X, et al.2025. Cycling Dof Factor 3 mediates light-dependent ascorbate biosynthesis by activating GDP-l-galactose phosphorylase in Rosa roxburghii fruit[J]. Plant Physiology, 197(1): 014.
[17] Jia D, Gao H, He Y, et al.2023. Kiwifruit monodehydroascorbate reductase 3 gene negatively regulates the accumulation of ascorbic acid in fruit of transgenic tomato plants[J]. International Journal of Molecular Sciences, 24(24): 17182.
[18] Liu X Y, Bulley S M, Varkonyi-Gasic E, et al.2023. Kiwifruit bZIP transcription factor AcePosF21 elicits ascorbic acid biosynthesis during cold stress[J]. Plant Physiology, 192(2): 982-999.
[19] Murre C, McCaw PS, Baltimore D.1989. A new DNA binding and dimerization motif in immunoglobulin enhancer binding,daughterless, MyoD,and myc proteins[J]. Cell, 56(5): 777-783.
[20] Mao K, Dong Q, Li C, et al.2017. Genome wide identification and characterization of apple bHLH transcription factors and expression analysis in response to drought and salt stress[J]. Frontiers in Plant Science, 8: 480.
[21] Niu X, Guan Y, Chen S, et al.2017. Genome-wide analysis of basic helix-loop-helix (bHLH) transcription factors in Brachypodium distachyon[J]. BMC Genomics, 18: 1-20.
[22] Song C B, Shan W, Kuang J F, et al.2020. The basic helix-loop-helix transcription factor MabHLH7 positively regulates cell wall-modifying-related genes during banana fruit ripening[J]. Postharvest Biology and Technology, 161: 111068.
[23] Shen T, Wen X, Wen Z, et al.2021. Genome-wide identification and expression analysis of bHLH transcription factor family in response to cold stress in sweet cherry (Prunus avium L.)[J]. Scientia Horticulturae, 279: 109905.
[24] Thoben C, Pucker B.2023. Automatic annotation of the bHLH gene family in plants[J]. BMC Genomics, 24: 780.
[25] Voronova A, Baltimore D.1990. Mutations that disrupt DNA binding and dimer formation in the E47 helix-loop-helix protein map to distinct domains[J]. Proceedings of the National Academy of Sciences of the USA, 87(12): 4722-4726.
[26] Waseem M, Li N, Su D, et al.2019. Overexpression of a basic helix-loop-helix transcription factor gene, SlbHLH22, promotes early flowering and accelerates fruit ripening in tomato (Solanum lycopersicum L.)[J]. Planta, 250: 173-185.
[27] Wang P, Su L, Gao H, et al.2018. Genome-wide characterization of bHLH genes in grape and analysis of their potential relevance to abiotic stress tolerance and secondary metabolite biosynthesis[J]. Frontiers in Plant Science, 9: 64.
[28] Wang W Y, Yu J Q, Du M C, et al.2022. Basic helix-loop-helix (bHLH) transcription factor MdbHLH3 negatively affects the storage performance of postharvest apple fruit[J]. Horticultural Plant Journal, 8(6): 700-712.
[29] Wu P, Li B, Liu Y, et al.2024. Multiple physiological and biochemical functions of ascorbic acid in plant growth, development,and abiotic stress response[J]. International Journal of Molecular Sciences, 25(3): 1832.
[30] Wu Y J, Xie M, Zhang Q C, et al.2009. Characteristics of 'White': A new easy-peel cultivar of Actinidia eriantha[J]. New Zealand Journal of Crop and Horticultural Science, 37(4): 369-373.
[31] Xing C, Liu Y, Zhao L, et al.2018. A novel MYB transcription factor regulates ascorbic acid synthesis and affects cold tolerance[J]. Plant Cell Environment, 42(3): 832-845.
[32] Yuan J, Yu Z, Lin T, et al.2019. BcERF070, a novel ERF (ethylene-response factor) transcription factor from non-heading Chinese cabbage, affects the accumulation of ascorbic acid by regulating ascorbic acid-related genes[J]. Molecular Breeding, 40(2): 85-94.
[33] Yao M M, Ge W Y, Zhou Q, et al.2021. Exogenous glutathione alleviates chilling injury in postharvest bell pepper by modulating the ascorbate-glutathione (AsA-GSH) cycle[J]. Food Chemistry, 352: 129458.
[34] Yang, Z, Lin L, Lu M, et al.2025. A bHLH transcription factor RrUNE12 regulates salt tolerance and promotes ascorbate synthesis[J]. Plant Cell Reports, 44: 42.
[35] Zhang D, Sun L, Xi D, et al.2024. Methyl jasmonate-induced bHLH42 mediates tissue-specific accumulation of anthocyanins via regulating flavonoid metabolism-related pathways in Caitai[J]. Physiologia Plantarum, 176(4): e14434.
[36] Zhang L C, Kang J, Xie Q L, et al.2020. The basic helix-loop-helix (bHLH) transcription factor SlbHLH95 affects fruit ripening and multiple metabolisms in tomato[J]. Journal of Experimental Botany, 71(20): 6311-6327.
[37] Zhang Y, Wang K, Xiao X, et al.2021. Effect of 1-MCP on the regulation processes involved in ascorbate metabolism in kiwifruit[J]. Postharvest Biology and Technology, 179: 111563.
[38] Zhao Q, Li J Q, Wang B T, et al.2024. Genome-wide analysis of the bHLH family and identification of bHLH genes involved in fruit development and ripening of cultivated octoploid strawberry[J]. Food Quality and Safety, 8: 14. |
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