基于红蓝光调控分析VvBES1-1参与‘红地球’葡萄花芽的分化

doi:10.3864/j.issn.0578-1752.2025.08.015

摘要/Abstract

摘要：

【目的】BRI1-EMS-Suppressor 1（BES1）是油菜素内酯（BR）信号传导的重要转录因子，调节植物光形态建成，并参与光周期开花。研究红光和蓝光调控下BES1在‘红地球’葡萄花芽分化中的功能，为解析红光和蓝光介导的BR调控机制奠定基础，也为研究其他木本果树的开花调控提供参考。【方法】应用生物信息学分析VvBES1-1蛋白结构及序列比对。以‘红地球’葡萄花芽为试材，以温室自然光为对照（CK），设置红蓝4﹕1（R4B1）为光照处理，采集‘红地球’葡萄花芽，利用qPCR分析VvBES1-1在‘红地球’葡萄花芽分化过程中的时空表达模式和组织特异性。运用转化烟草方法进行亚细胞定位。通过双分子荧光互补（BiFC）试验探究VvBES1-1与其他蛋白的相互作用，以及运用酵母自激活活性测定评估VvBES1-1的转录激活能力。【结果】VvBES1-1具有BES1_N结构域，是一种BES1-S型蛋白，与毛果杨BES1亲缘关系最近，在‘红地球’葡萄各时期均有表达。在R4B1处理期间，与对照相比，VvBES1-1表达量显著降低。在花序次轴发育期，VvBES1-1表达量达到最高，此外，经10 mmol·L^-1EBR处理后，促进了‘红地球’葡萄花芽VvBES1-1的表达。酵母自激活试验表明，VvBES1-1具有自激活活性。亚细胞定位结果表明，VvBES1-1定位于细胞核。过表达VvBES1-1烟草延迟开花时间，促进茎的伸长，并增加了分生组织的数量。VvBES1-1的下调表达则整合了BR和光周期信号，促进了‘红地球’葡萄花芽的分化。在‘红地球’葡萄花芽分化过程中，VvBES1-1受红光和蓝光的显著调节，且在R4B1处理下，VvBES1-1的表达水平在10 mmol·L^-1的EBR处理6 h时达到最高。【结论】VvBES1-1在‘红地球’葡萄花芽分化中起重要作用。VvBES-1能够整合BR与光周期信号抑制葡萄花芽分化。而在红蓝光的影响下，能够有效下调VvBES-1的表达，达到促进葡萄花芽分化的目的。

关键词: ‘红地球’葡萄, VvBES1-1, 红蓝光, BR, 花芽分化

Abstract:

【Objective】 BRI1-EMS-Suppressor 1 (BES1), a key transcription factor in brassinosteroid (BR) signaling, regulates plant photomorphogenesis and photoperiodic flowering. This study aimed to investigate the role of BES1 in flower bud differentiation of Vitis vinifera Red Globe under red and blue light regulation, thereby elucidating the BR-mediated mechanisms driven by light quality and providing insights into flowering regulation in other woody fruit trees.【Method】 Bioinformatics analysis was performed to characterize the protein structure and sequence alignment of VvBES1-1. Flower buds of Red Globe grape were collected under greenhouse natural light (CK, control) and red:blue=4:1 (R4B1) light treatment. qPCR was used to analyze the spatiotemporal expression patterns and tissue specificity of VvBES1-1 during flower bud differentiation. Subcellular localization was determined via tobacco (Nicotiana benthamiana) transient transformation. Protein-protein interactions were examined using bimolecular fluorescence complementation (BiFC), and transcriptional activation activity was assessed via yeast autoactivation assays. 【Result】VvBES1-1 contains a BES1_N domain and belongs to the BES1-S-type protein family, showing the closest phylogenetic relationship with Populus trichocarpa BES1. It is expressed throughout all developmental stages of Red Globe grapevines. During R4B1 treatment, the expression level of VvBES1-1 was significantly reduced compared to the control. Its expression peaked during the development of secondary inflorescence axes. Additionally, treatment with 10 mmol·L^-¹ EBR enhanced VvBES1-1 expression in Red Globe grape flower buds. Yeast autoactivation assays demonstrated that VvBES1-1 possesses self-activation activity. Subcellular localization analysis revealed that VvBES1-1 is localized in the nucleus. Overexpression of VvBES1-1 in tobacco delayed flowering time, promoted stem elongation, and increased meristem number. Downregulation of VvBES1-1 expression integrated brassinosteroid (BR) and photoperiod signaling pathways to promote flower bud differentiation in Red Globe grapes. During flower bud differentiation, VvBES1-1 expression was significantly regulated by red and blue light. Under R4B1 treatment, its expression peaked after 6 hours of 10 mmol·L^-¹ EBR exposure.【Conclusion】VvBES1-1 plays a critical role in flower bud differentiation of Red Globe grapes. It integrates BR and photoperiod signals to inhibit grape flower bud differentiation. However, under red and blue light conditions, the expression of VvBES1-1 is effectively downregulated, thereby promoting flower bud differentiation. This study provides insights into the regulatory mechanisms of light and phytohormones in grape reproductive development.

Key words: Red Globe grape, VvBES1-1, red and blue light, BR, bud differentiation

汤学燊, 党仕卓, 周娟, 李佳豪, 李梅花, 胡豪, 张亚红. 基于红蓝光调控分析VvBES1-1参与‘红地球’葡萄花芽的分化[J]. 中国农业科学, 2025, 58(8): 1650-1662.

TANG XueShen, DANG ShiZhuo, ZHOU Juan, LI JiaHao, LI MeiHua, HU Hao, ZHANG YaHong. Analysis of VvBES1-1 Involvement in Flower Bud Differentiation of Red Globe Grape Based on Red and Blue Light Regulation[J]. Scientia Agricultura Sinica, 2025, 58(8): 1650-1662.

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参考文献 48

[1]	JAILLON O, AURY J, NOEL B, POLICRITI A, CLÉPET C, CASAGRANDE A, CHOISNE N, AUBOURG S, VITULO N, JUBIN C, et al. The grapevine genome sequence suggests ancestral hexaploidization in major angiosperm Phyla. Nature, 2007, 449: 463-467.
[2]	NORIEGA X, PÉREZ F J. Cell cycle genes are activated earlier than respiratory genes during release of grapevine buds from endodormancy. Plant Signaling & Behavior, 2017, 12(10): e1321189.
[3]	VICTORINO G F, BRAGA R, SANTOS-VICTOR J, LOPES C M. Yield components detection and image-based indicators for non-invasive grapevine yield prediction at different phenological phases. OENO ONE, 2020, 54(4): 833-848.
[4]	WAN S B, LI W L, ZHU Y Y, LIU Z M, HUANG W D, ZHAN J C. Genome-wide identification, characterization and expression analysis of the auxin response factor gene family in Vitis vinifera. Plant Cell Reports, 2014, 33(8): 1365-1375. doi: 10.1007/s00299-014-1622-7 pmid: 24792421
[5]	LI-MALLET A, RABOT A, GENY L. Factors controlling inflorescence primordia formation of grapevine: Their role in latent bud fruitfulness? A review. Botany, 2016, 94(3): 147-163.
[6]	BOSS P K, THOMAS M R. Association of dwarfism and floral induction with a grape green revolution mutation. Nature, 2002, 416(6883): 847-850.
[7]	LIU X, YUAN M, DANG S Z, ZHOU J, ZHANG Y H. Comparative transcriptomic analysis of transcription factors and hormones during flower bud differentiation in ‘Red Globe’ grape under red-blue light. Scientific Reports, 2023, 13(1): 8932.
[8]	CARMONA M J, CHAÏB J, MARTÍNEZ-ZAPATER J M, THOMAS M R. A molecular genetic perspective of reproductive development in grapevine. Journal of Experimental Botany, 2008, 59(10): 2579-2596. doi: 10.1093/jxb/ern160 pmid: 18596111
[9]	CACKETT L, LUGINBUEHL L H, SCHREIER T B, LOPEZ-JUEZ E, HIBBERD J M. Chloroplast development in green plant tissues: The interplay between light, hormone, and transcriptional regulation. New Phytologist, 2022, 233(5): 2000-2016.
[10]	LAVEE S. Necrosis in grapevine buds (Vitis vinifera cv. Queen of Vineyard) III. Endogenous gibberellin levels in leaves and buds. Vitis: Journal of Grapevine Research, 2015, 26: 225.
[11]	CHEN Y Q, TAI S S, WANG D W, DING A M, SUN T T, WANG W F, SUN Y H. Homology-based analysis of the GRAS gene family in tobacco. Genetics and Molecular Research, 2015, 14(4): 15188-15200. doi: 10.4238/2015.November.25.7 pmid: 26634482
[12]	DE WIT M, GALVÃO V C, FANKHAUSER C. Light-mediated hormonal regulation of plant growth and development. Annual Review of Plant Biology, 2016, 67: 513-537. doi: 10.1146/annurev-arplant-043015-112252 pmid: 26905653
[13]	SU D D, XIANG W, LIANG Q, WEN L, SHI Y, SONG B Q, LIU Y D, XIAN Z Q, LI Z G. Tomato SlBES1.8 influences leaf morphogenesis by mediating gibberellin metabolism and signaling. Plant & Cell Physiology, 2022, 63(4): 535-549.
[14]	LIU Y, JAFARI F, WANG H Y. Integration of light and hormone signaling pathways in the regulation of plant shade avoidance syndrome. aBIOTECH, 2021, 2(2): 131-145. doi: 10.1007/s42994-021-00038-1 pmid: 36304753
[15]	LI J, TERZAGHI W, GONG Y Y, LI C R, LING J J, FAN Y Y, QIN N X, GONG X Q, ZHU D M, DENG X W. Modulation of BIN2 kinase activity by HY5 controls hypocotyl elongation in the light. Nature Communications, 2020, 11(1): 1592. doi: 10.1038/s41467-020-15394-7 pmid: 32221308
[16]	ZHU J, JUTHAMAS S S, WANG Z. Brassinosteroid signalling. Development, 2013, 140: 1615-1620.
[17]	XIA X J, FANG P P, GUO X, QIAN X J, ZHOU J, SHI K, ZHOU Y H, YU J Q. Brassinosteroid-mediated apoplastic H₂O₂-glutaredoxin 12/14 cascade regulates antioxidant capacity in response to chilling in tomato. Plant, Cell & Environment, 2018, 41(5): 1052-1064.
[18]	SYMONS G M, DAVIES C, SHAVRUKOV Y, DRY I B, REID J B, THOMAS M R. Grapes on steroids. Brassinosteroids are involved in grape berry ripening. Plant Physiology, 2006, 140(1): 150-158. doi: 10.1104/pp.105.070706 pmid: 16361521
[19]	陈慧泽, 刁丽曼, 周佳佳, 韩榕, 杜美婷. 油菜素内酯与其他植物激素互作调控植物生长发育及胁迫响应的研究进展. 植物研究, 2024, 44(6): 812-821. doi: 10.7525/j.issn.1673-5102.2024.06.002
	CHEN H Z, DIAO L M, ZHOU J J, HAN R, DU M T. Research advances in brassinosteroids interaction with other phytohormones regulating plant growth, development and stress responses. Bulletin of Botanical Research, 2024, 44(6): 812-821. (in Chinese)
[20]	RUAN J X, CHEN H H, ZHU T, YU Y G, LEI Y W, YUAN L B, LIU J, WANG Z Y, KUANG J F, LU W J, HUANG S Z, LI C L. Brassinosteroids repress the seed maturation program during the seed-to-seedling transition. Plant Physiology, 2021, 186(1): 534-548. doi: 10.1093/plphys/kiab089 pmid: 33620498
[21]	SHIGETA T, ZAIZEN Y, SUGIMOTO Y, NAKAMURA Y, MATSUO T, OKAMOTO S. Heat shock protein 90 acts in brassinosteroid signaling through interaction with BES1/BZR1 transcription factor. Journal of Plant Physiology, 2015, 178: 69-73. doi: 10.1016/j.jplph.2015.02.003 pmid: 25778412
[22]	CHEN W Y, LV M H, WANG Y Z, WANG P G, CUI Y W, LI M Z, WANG R S, GOU X P, LI J. BES1 is activated by EMS1-TPD1- SERK1/2-mediated signaling to control tapetum development in Arabidopsis thaliana. Nature Communications, 2019, 10(1): 4164.
[23]	CHENG P L, LIU Y N, YANG Y M, CHEN H, CHENG H, HU Q, ZHANG Z X, GAO J J, ZHANG J X, DING L, FANG W M, CHEN S M, CHEN F D, JIANG J F. CmBES1 is a regulator of boundary formation in Chrysanthemum ray florets. Horticulture Research, 2020, 7(1): 129.
[24]	MECCHIA M A, GARCÍA-HOURQUET M, LOZANO-ELENA F, PLANAS-RIVEROLA A, BLASCO-ESCAMEZ D, MARQUÈS- BUENO M, MORA-GARCÍA S, CAÑO-DELGADO A I. The BES1/BZR1-family transcription factor MpBES1 regulates cell division and differentiation in Marchantia polymorpha. Current Biology, 2021, 31(21): 4860-4869.e8.
[25]	MARTÍNEZ C, ESPINOSA-RUÍZ A, DE LUCAS M, BERNARDO- GARCÍA S, FRANCO-ZORRILLA J M, PRAT S. PIF4-induced BR synthesis is critical to diurnal and thermomorphogenic growth. The EMBO Journal, 2018, 37(23): e99552.
[26]	WANG F, GAO Y S, LIU Y W, ZHANG X, GU X X, MA D B, ZHAO Z W, YUAN Z J, XUE H W, LIU H T. BES1-regulated BEE1 controls photoperiodic flowering downstream of blue light signaling pathway in Arabidopsis. New Phytologist, 2019, 223(3): 1407-1419.
[27]	FELLER A, MACHEMER K, BRAUN E L, GROTEWOLD E. Evolutionary and comparative analysis of MYB and bHLH plant transcription factors. The Plant Journal, 2011, 66(1): 94-116. doi: 10.1111/j.1365-313X.2010.04459.x pmid: 21443626
[28]	TOLEDO-ORTIZ G, HUQ E, QUAIL P H. The Arabidopsis basic/ helix-loop-helix transcription factor family. The Plant Cell, 2003, 15(8): 1749-1770.
[29]	FRIEDRICHSEN D M, NEMHAUSER J, MURAMITSU T, MALOOF J N, ALONSO J, ECKER J R, FURUYA M, CHORY J. Three redundant brassinosteroid early response genes encode putative bHLH transcription factors required for normal growth. Genetics, 2002, 162(3): 1445-1456. doi: 10.1093/genetics/162.3.1445 pmid: 12454087
[30]	LEE M H, KIM B, SONG S K, HEO J O, YU N N, LEE S A, KIM M, KIM D G, SOHN S O, LIM C E, CHANG K S, LEE M M, LIM J. Large-scale analysis of the GRAS gene family in Arabidopsis thaliana. Plant Molecular Biology, 2008, 67(6): 659-670.
[31]	SONG X M, LIU T K, DUAN W K, MA Q H, REN J, WANG Z, LI Y, HOU X L. Genome-wide analysis of the GRAS gene family in Chinese cabbage (Brassica rapa ssp. pekinensis). Genomics, 2014, 103(1): 135-146.
[32]	FANG P P, YAN M Y, CHI C, WANG M Q, ZHOU Y H, ZHOU J, SHI K, XIA X J, FOYER C H, YU J Q. Brassinosteroids act as a positive regulator of photoprotection in response to chilling stress. Plant Physiology, 2019, 180(4): 2061-2076. doi: 10.1104/pp.19.00088 pmid: 31189657
[33]	SYMONS G M, REID J B. Hormone levels and response during de-etiolation in pea. Planta, 2003, 216(3): 422-431. doi: 10.1007/s00425-002-0860-z pmid: 12520333
[34]	LI J M, CHORY J. A putative leucine-rich repeat receptor kinase involved in brassinosteroid signal transduction. Cell, 1997, 90(5): 929-938. doi: 10.1016/s0092-8674(00)80357-8 pmid: 9298904
[35]	JIANG J J, ZHANG C, WANG X L. A recently evolved isoform of the transcription factor BES1 promotes brassinosteroid signaling and development in Arabidopsis thaliana. The Plant Cell, 2015, 27(2): 361-374.
[36]	LI J M, NAM K H. Regulation of brassinosteroid signaling by a GSK3/SHAGGY-like kinase. Science, 2002, 295(5558): 1299-1301. doi: 10.1126/science.1065769 pmid: 11847343
[37]	ZHAO J, PENG P, SCHMITZ R J, DECKER A D, TAX F E, LI J M. Two putative BIN2 substrates are nuclear components of brassinosteroid signaling. Plant Physiology, 2002, 130(3): 1221-1229. doi: 10.1104/pp.102.010918 pmid: 12427989
[38]	VERT G, CHORY J. Downstream nuclear events in brassinosteroid signalling. Nature, 2006, 441(7089): 96-100.
[39]	WANG Z Y, NAKANO T, GENDRON J, HE J X, CHEN M, VAFEADOS D, YANG Y L, FUJIOKA S, YOSHIDA S, ASAMI T, CHORY J. Nuclear-localized BZR1 mediates brassinosteroid-induced growth and feedback suppression of brassinosteroid biosynthesis. Developmental Cell, 2002, 2(4): 505-513. doi: 10.1016/s1534-5807(02)00153-3 pmid: 11970900
[40]	LI J J, WANG L, LENG F, MA C, ZHANG C X, WANG S P. Genome-wide identification, characterization and gene expression of BES1 transcription factor family in grapevine (Vitis vinifera L.). Scientific Reports, 2023, 13(1): 240.
[41]	LIU Z, QANMBER G, LU L L, QIN W Q, LIU J, LI J, MA S Y, YANG Z E, YANG Z R. Genome-wide analysis of BES1 genes in Gossypium revealed their evolutionary conserved roles in brassinosteroid signaling. Science China Life Sciences, 2018, 61(12): 1566-1582.
[42]	GAO A J, WENG W F, YAO X, WU W J, BAI Q, XIONG R Q, MA C, CHENG J P, RUAN J J. Genome-wide identification, structural characterization, and gene expression analysis of BES1 transcription factor family in Tartary buckwheat (Fagopyrum tataricum). Agronomy, 2022, 12(11): 2729.
[43]	CAO X, KHALIQ A, LU S, XIE M, MA Z, MAO J, CHEN B. Genome-wide identification and characterization of the BES1 gene family in apple (Malus domestica). Plant Biology, 2020, 22(4): 723-733. doi: 10.1111/plb.13109 pmid: 32141196
[44]	CAO X J, MA W F, ZENG F W, CHENG Y J, MA Z H, MAO J, CHEN B H. Grape BES1 transcription factor gene VvBES1-3 confers salt tolerance in transgenic Arabidopsis. Gene, 2023, 854: 147059.
[45]	HAMASAKI H, AYANO M, NAKAMURA A, FUJIOKA S, ASAMI T, TAKATSUTO S, YOSHIDA S, OKA Y, MATSUI M, SHIMADA Y. Light activates brassinosteroid biosynthesis to promote hook opening and petiole development in Arabidopsis thaliana. Plant & Cell Physiology, 2020, 61(7): 1239-1251.
[46]	郭飞梅, 吕铭辉, 黎家. 油菜素甾醇的稳态与信号转导调控研究进展. 植物生理学报, 2023, 59(12): 2217-2240.
	GUO F M, LÜ M H, LI J. research advances in the homeostasis and signal transduction regulation of brassinosteroids. Plant Physiology Journal, 2023, 59(12): 2217-2240. (in Chinese)
[47]	CONWAY S J, WALCHER-CHEVILLET C L, SALOME BARBOUR K, KRAMER E M. Brassinosteroids regulate petal spur length in Aquilegia by controlling cell elongation. Annals of Botany, 2021, 128(7): 931-942.
[48]	YU H Q, FENG W Q, SUN F A, ZHANG Y Y, QU J T, LIU B L, LU F Z, YANG L, FU F L, LI W C. Cloning and characterization of BES1/BZR1 transcription factor genes in maize. Plant Growth Regulation, 2018, 86(2): 235-249.

功能 Function	引物名称 Primer name	引物序列 Primer sequence (5′-3′)
编码区（CDS）扩增 Coding region (CDS) amplification	VvBES1-1-1-pCR-F	ATGACAGGAACGAGGCTCCC
编码区（CDS）扩增 Coding region (CDS) amplification	VvBES1-1-pCR-R	TCTGGTTCTAGAGCTCCCAAGG
亚细胞定位载体构建 Subcellular localization vector construction	VvBES1-221-F	ATGACAGGAACGAGGCTCCCAACATG
亚细胞定位载体构建 Subcellular localization vector construction	VvBES1-221-R	TCTGGTTCTAGAGCTCCCAAGG
酵母自激活载体构建 Yeast self-activating vector construction	VvBES1-BD-F	ATGACAGGAACGAGGCTCCC
酵母自激活载体构建 Yeast self-activating vector construction	VvBES1-BD-R	TCTGGTTCTAGAGCTCCCAAGG
葡萄内参基因 Internal reference genes of grape	VvActin-F	GATTCTGGTGATGGTGTGAGT
葡萄内参基因 Internal reference genes of grape	VvActin-R	GACAATTTCCCGTTCAGCAGT
VvBES1表达模式分析（葡萄） VvBES1 expression pattern (Vitis vinifera)	VvBES1-1-F	TGGACGGTTGAAGAAGACGG
	VvBES1-1-R	TGGGCTTGGCTGGTATGATG
烟草荧光定量内参基因 Internal reference genes of tobacco	NT-Actin-F	GGCTTACATTGCTCTTGACTATGAAC
烟草荧光定量内参基因 Internal reference genes of tobacco	NT-Actin-R	ATCAGGCAGCTCTGTAGCTCTCTCT
VvBES1表达模式分析（烟草） VvBES1 Expression Pattern (Tobacco)	VvBES1-1-NT-F	GCTAGTGCTGATGCCAATGC
VvBES1表达模式分析（烟草） VvBES1 Expression Pattern (Tobacco)	VvBES1-1-NT-R	GTGGGGGAACTTGGATGAGG