Structural and Functional Analysis of Gibberellin Signaling Transduction Pathway Gene HbRGA1 in Rubber Tree

doi:10.7525/j.issn.1673-5102.2021.04.008

Abstract

Abstract:

DELLA protein is a negative regulator of gibberellin signaling pathway， which inhibits plant growth and development. Analyzing the structure and function of DELLA family members will be helpful to elucidate the mechanism by which the DELLA protein regulates the growth and development of rubber trees. In this study， the full-length cDNA sequence of HbRGA1 was cloned from the leaves of rubber tree cultivar Reyan73397. HbRGA1 is 2 136 bp long， contains 1 839 bp ORF， and encodes 613 amino acids. The HbRGA1 protein sequence contains DELLA and GRAS superfamily conserved domains， which are 82.5% like the DELLA genes of poplar， cassava and rubber trees. qRT-PCR analysis showed that the expression level of HbRGA1 was high in the leaf of rubber tree， but very low in the bark and latex. The expression of HbRGA1 in leaves was significantly up-regulated by gibberellin and abscisic acid spraying. These results indicated that HbRGA1 was closely related to gibberellin and other hormone signals in rubber trees， which laid a good foundation for further study of HbRGA1’s structure and function in the growth and development of rubber trees.

Key words: Hevea brasiliensis, HbRGA1, gibberellin, gene expression

CLC Number:

S794.1

Hua-Feng CHEN, Song-Le FAN, Li-Feng WANG. Structural and Functional Analysis of Gibberellin Signaling Transduction Pathway Gene HbRGA1 in Rubber Tree[J]. Bulletin of Botanical Research, 2021, 41(4): 531-539.

Figures/Tables 7

Table 1

Fig.1

Fig.2

Fig.3

Table 2

Fig.4

Fig.5

References 40

1	白云赫，朱旭东，樊秀彩，等.植物DELLA蛋白及其应答赤霉素信号调控植物生长发育的研究进展［J］.分子植物育种，2019，17（8）：2509-2516.
	Bai Y H，Zhu X D，Fan X C，et al.Reseach progress of plant DELLA proteins and its response to gibberellin signal regulating plant growth and development［J］.Molecular Plant Breeding，2019，17（8）：2509-2516.
2	Blanco-Touriñán N，Serrano-Mislata A，Alabadi D.Regulation of DELLA proteins by post-translational modifications［J］.Plant and Cell Physiology，2020，61（11）：1891-1901.
3	Phokas A，Coates J C.Evolution of DELLA function and signaling in land plants［J］.Evolution & Development，2021：e12365，doi：10.1111/ede.12365.
4	高秀华，傅向东.赤霉素信号转导及其调控植物生长发育的研究进展［J］.生物技术通报，2018，34（7）：1-13.
	Gao X H，Fu X D.Research progress for the gibberellin signaling and action on plant growth and development［J］.Biotechnology Bulletin，2018，34（7）：1-13.
5	Huang H，Gong Y，Liu B，et al.The DELLA proteins interact with MYB21 and MYB24 to regulate filament elongation in Arabidopsis［J］.BMC Plant Biology，2020，20（1）：64.
6	Yan J D，Li X M，Zeng B J，et al.FKF1 F-box protein promotes flowering in part by negatively regulating DELLA protein stability under long-day photoperiod in Arabidopsis［J］.Journal of Integrative Plant Biology，2020，62（11）：1717-1740.
7	Wu J J，Yan G B，Duan Z Q，et al.Roles of the Brassica napus DELLA Protein BnaA6.RGA，in modulating drought tolerance by interacting with the ABA signaling component BnaA10.ABF2［J］.Frontiers in Plant Science，2020，11：577.
8	卢艳芬，卜芋芬，郝素晓，等.海棠‘比利时垂枝’McRGL1a基因的克隆与分析［J］.北京农学院学报，2016，31（3）：1-5.
	Lu Y F，Bu Y F，Hao S X，et al.Cloning and analysis of McRGL1a in Malus sp.‘Belgium weeping’［J］.Journal of Beijing University of Agriculture，2016，31（3）：1-5.
9	麻楠，刘静，陈琦，等.苹果砧木‘SH6’中RGLs基因的克隆及生物信息学分析［J］.北京农学院学报，2018，33（2）：1-6.
	Ma N，Liu J，Chen Q，et al.Cloning and bioinformatic analysis of RGLs in apple rootstock ‘SH6’［J］.Journal of Beijing University of Agriculture，2018，33（2）：1-6.
10	黄昊，宫铭，殳晓强，等.珍珠黄杨BsGAI1基因的克隆及实时定量表达分析［J］.分子植物育种，2016，14（11）：3033-3040.
	Huang H，Gong M，Shu X Q，et al.Cloning and real-time quantitative expression analysis on BsGAI1 gene from Buxus sinica var. parvifolia［J］.Molecular Plant Breeding，2016，14（11）：3033-3040.
11	李晓晨，李国瑞，丛安琪，等.蓖麻RcDella（GAI）基因克隆及生物信息学分析［J］.分子植物育种，2018，16（20）：6598-6603.
	Li X C，Li G R，Cong A Q，et al.Cloning and bioinformatics analysis of RcDella（GAI） gene in castor［J］.Molecular Plant Breeding，2018，16（20）：6598-6603.
12	何长辉，莫业勇，刘锐金.中国天然橡胶生产能力预测分析（2019~2025年）［J］.林业经济问题，2020，40（3）：320-327.
	He C H，Mo Y Y，Liu R J.Forecasting of natural rubber production capacity in China（2019－2025）［J］.Issues of Forestry Economics，2020，40（3）：320-327.
13	黄珍珠，李寅，陈慧华，等.基于气象关键因子的广东省橡胶产量预报［J］.热带农业科学，2018，38（2）：107-112.
	Huang Z Z，Li Y，Chen H H，et al.Forecast of rubber production based on meteorological key factors in Guangdong province［J］.Chinese Journal of Tropical Agriculture，2018，38（2）：107-112.
14	Chao J，Huang Z J，Yang S G，et al.Genome-wide identification and expression analysis of the phosphatase 2A family in rubber tree（Hevea brasiliensis）［J］.PLoS One，2020，15（2）：e0228219.
15	张冬，李晓娜，肖厚贞，等.巴西橡胶树HbbZIP40转录因子基因的克隆与分析［J］.基因组学与应用生物学，2018，37（9）：3926-3932.
	Zhang D，Li X N，Xiao H Z，et al.Molecular cloning and characterization of transcription factor gene HbbZIP40 in rubber tree（Hevea brasiliensis Muell.Arg.）［J］.Genomics and Applied Biology，2018，37（9）：3926-3932.
16	李晓娜，肖厚贞，万三连，等.巴西橡胶树生长素响应HbJAR1基因克隆与表达分析［J］.热带作物学报，2017，38（8）：1478-1484.
	Li X N，Xiao H Z，Wan S L，et al.Gene cloning and expression analysis of auxin responsive HbJAR1 in rubber tree［J］.Chinese Journal of Tropical Crops，2017，38（8）：1478-1484.
17	Piyatrakul P，Putranto R A，Martin F，et al.Some ethylene biosynthesis and AP2/ERF genes reveal a specific pattern of expression during somatic embryogenesis in Hevea brasiliensis［J］.BMC Plant Biology，2012，12（1）：244.
18	Chen L G，Xiang S Y，Chen Y L，et al.Arabidopsis WRKY45 interacts with the DELLA protein RGL1 to positively regulate age-triggered leaf senescence［J］.Molecular Plant，2017，10（9）：1174-1189.
19	Wang L F.Physiological and molecular responses to drought stress in rubber tree（Hevea brasiliensis Muell.Arg.）［J］.Plant Physiology and Biochemistry，2014，83：243-249.
20	Wang L F，Wang J K，An F，et al.Molecular cloning and characterization of a stress responsive peroxidase gene HbPRX42 from rubber tree［J］.Brazilian Journal of Botany，2016，39（2）：475-483.
21	陆燕茜，张冬，王立丰，等.巴西橡胶树HbMYB62转录因子基因的克隆和表达分析［J］.植物研究，2017，37（6）：953-960，969.
	Lu Y X，Zhang D，Wang L F，et al.Cloning and expressional analysis of HbMYB62 under multi-stimulation in Hevea brasiliensis Muell.Arg［J］．Bulletin of Botanical Research，2017，37（6）：953-960，969.
22	Dill A，Jung H S，Sun T P.The DELLA motif is essential for gibberellin-induced degradation of RGA［J］.Proceedings of the National Academy of Sciences of the United States of America，2001，98（24）：14162-14167.
23	Itoh H，Shimada A，Ueguchi-Tanaka M，et al.Over expression of a GRAS protein lacking the DELLA domain confers altered gibberellin responses in rice［J］.The Plant Journal，2005，44（4）：669-679.
24	Sato T，Miyanoiri Y，Takeda M，et al.Expression and purification of a GRAS domain of SLR1，the rice DELLA protein［J］.Protein Expression and Purification，2014，95：248-258.
25	Sheerin D J，Buchanan J，Kirk C，et al.Inter- and intra-molecular interactions of Arabidopsis thaliana DELLA protein RGL1［J］.Biochemical Journal，2011，435（3）：629-639.
26	Fu X D，Richards D E，Ait-Ali T，et al.Gibberellin-mediated proteasome-dependent degradation of the barley DELLA protein SLN1 repressor［J］.The Plant Cell，2002，14（12）：3191-3200.
27	Lee S，Cheng H，King K E，et al.Gibberellin regulates Arabidopsis seed germination via RGL2，a GAI/RGA-like gene whose expression is up-regulated following imbibition［J］.Genes & Development，2002，16（5）：646-658.
28	Achard P，Vriezen W H，Van Der Straeten D，et al.Ethylene regulates Arabidopsis development via the modulation of DELLA protein growth repressor function［J］.The Plant Cell，2003，15（12）：2816-2825.
29	Reid J B，Davidson S E，Ross J J.Auxin acts independently of DELLA proteins in regulating gibberellin levels［J］.Plant Signaling & Behavior，2011，6（3）：406-408.
30	Hou X L，Lee L Y C，Xia K F，et al.DELLAs modulate jasmonate signaling via competitive binding to JAZs［J］.Developmental Cell，2010，19（6）：884-894.
31	Achard P，Gong F，Cheminant S，et al.The cold-inducible CBF1 factor-dependent signaling pathway modulates the accumulation of the growth-repressing DELLA proteins via its effect on gibberellin metabolism［J］.The Plant Cell，2008，20（8）：2117-2129.
32	Piskurewicz U，Turečková V，Lacombe E，et al.Far-red light inhibits germination through DELLA-dependent stimulation of ABA synthesis and ABI3 activity［J］.The EMBO Journal，2009，28（15）：2259-2271.
33	吴绍华，张世鑫，陈月异，等.橡胶树DELLA蛋白编码基因HbGAI的克隆与表达分析［J］.西北植物学报，2015，35（11）：2157-2163.
	Wu S H，Zhang S X，Chen Y Y，et al.Cloning and expression analysis of HbGAI gene in rubber tree（Hevea brasiliensis Muell.Arg.）［J］.Acta Botanica Boreali-Occidentalia Sinica，2015，35（11）：2157-2163.
34	陈宇杰，刘飞，梁菲菲，等.蓖麻DELLA蛋白家族GAI基因克隆、表达及生物信息学分析［J］.内蒙古民族大学学报：自然科学版，2017，32（4）：320-327.
	Chen Y J，Liu F，Liang F F，et al.Cloning of castor DELLA protein family GAI gene expression and analysis of bioinformatics［J］.Journal of Inner Mongolia University for Nationalities，2017，32（4）：320-327.
35	Tomlinson L，Yang Y，Emenecker R，et al.Using CRISPR/Cas9 genome editing in tomato to create a gibberellin-responsive dominant dwarf DELLA allele［J］.Plant Biotechnology Journal，2019，17（1）：132-140.
36	Noël L D，Cagna G，Stuttmann J，et al.Interaction between SGT1 and cytosolic/nuclear HSC70 chaperones regulates Arabidopsis immune responses［J］.The Plant Cell，2007，19（12）：4061-4076.
37	Chao J Q，Chen Y Y，Wu S H，et al.Comparative transcriptome analysis of latex from rubber tree clone CATAS8-79 and PR107 reveals new cues for the regulation of latex regeneration and duration of latex flow［J］.BMC Plant Biology，2015，15（1）：104.
38	Chao J Q，Yang S G，Chen Y Y，et al.Evaluation of reference genes for quantitative real-time PCR analysis of the gene expression in laticifers on the basis of latex flow in rubber tree（Hevea brasiliensis Muell.Arg.）［J］.Frontiers in Plant Science，2016，7：1149.
39	Osborne D J，Sargent J A.A model for the mechanism of stimulation of latex flow in Hevea brasiliensis by ethylene［J］.Annals of Applied Biology，1974，78（1）：83-88.
40	Shi M J，Cai F G，Tian W M.Ethrel-stimulated prolongation of latex flow in the rubber tree（Hevea brasiliensis Muell.Arg.）：an Hev b 7-like protein acts as a universal antagonist of rubber particle aggregating factors from lutoids and C-serum［J］.The Journal of Biochemistry，2016，159（2）：209-216.

引物名称 Primer name	引物序列 Primer sequence(5′—3′)
HbRGA1 F	GAAAGATGAAAAGAGATCACCAAGA
HbRGA1 R	ACTGACTCGCTAACATTAACTCACC
HbRGA1 QF	CCTCTCTTAGCTGATGCTT
HbRGA1 QR	GCGAGTCAATTAAGACAACC
HbACTIN F (Genbank：HQ260674.1)	GATGTGGATATCAGGAAGGA
HbACTIN R	CATACTGCTTGGAGCAAGA

项目 Project	理化性质 Physicochemical property
分子式Formula	C₂₉₁₈H₄₅₉₁N₈₀₃O₉₁₄S₂₉
相对分子质量Molecular weight (kDa)	66476.12
等电点Theoretical pI	5.19
正电荷残基数 Number of positive charge residues (Arg+Lys)	49
负电荷残基数 Number of negatively charge residues(Asp+Glu)	71
不稳定系数Instability index	50.01（大于40为不稳定）
脂肪系数Aliphatic index	82.94
总平均亲水性 Grand average of hydropathicity (GRAVY)	-0.220

[1]	Siqi WANG, Yunjie BI, Huixian LI, Zhengbao SONG, Runke LIU, Ruixin GAO. Effect of Exogenous Gibberellin on Seed Germination of Agastache rugosa under Salt Stress [J]. Bulletin of Botanical Research, 2026, 46(3): 481-492.
[2]	Banghuan WANG, Jiaxin LI, Qi PAN, Yueqin LIANG, Aiping WU, Shuangjiao FENG, Ju GU, Zixiang YANG, Ping LIU, Chao WANG. Tissue Structure, Hormone Levels, and Molecular Changes in Horned Galls Before and After Dehiscence [J]. Bulletin of Botanical Research, 2026, 46(3): 493-504.
[3]	Wenbin DING, Yuehan HU, Yinjia BAI, Yuhan YANG, Hua XUE. Expression Characteristics and Functional Analysis of PtNF-Ys Gene during Early Flower Development in Pinus tabuliformis [J]. Bulletin of Botanical Research, 2026, 46(2): 195-208.
[4]	Mengmeng CHENG, Bin GE, Beisen KOU, Wenbin GUAN, Hai LU, Huihong GUO, Hui LI. Identification and Expression Analysis of HDAC and HAT gene families in Xanthoceras sorbifolium [J]. Bulletin of Botanical Research, 2026, 46(1): 51-66.
[5]	Chengming XIONG, Qiaoxin WANG, Liming HE, Jianfei LIU, Guangzhou PENG, Anhong HU, Xinyu LI, Yaguang ZHAN. Impact of Rejuvenation Measures on Growth and Tissue Culture Regeneration of Fraxinus mandshurica [J]. Bulletin of Botanical Research, 2026, 46(1): 91-100.
[6]	Qingqing XU, Dehui SUN, Wenmeng WANG, Dangdang LI, Xingguo LAN, Guangchao SUI. Research Progress of Plant Growth, Development and Stress Resistance Traits Regulated by Liquid-liquid Phase Separation-mediated Gene Expression [J]. Bulletin of Botanical Research, 2024, 44(6): 805-811.
[7]	Bingbing GUO, Mingyang LIU, Longjun DAI, Hong YANG, Lifeng WANG. Research Progress on the Mechanism of Plant Hormone Regulating Yield and Latex Flow in Hevea brasiliensis [J]. Bulletin of Botanical Research, 2024, 44(2): 161-167.
[8]	Huiling YAN, Xinxin ZHANG, Xiyang ZHAO, Guanzheng QU, Zhaoning WANG, Rui HAN. Effects of Light Signal on Anthocyanin Biosynthesis and Gene Expression of AmRosea1 Overexpressed 84K Poplar [J]. Bulletin of Botanical Research, 2023, 43(6): 815-825.
[9]	Lei XU, Xiao XU, Qinsong LIU. Effects of Exogenous Salicylic Acid on Antioxidant System and Gene Expression of Davidia involucrata Seedlings under Salt Stress [J]. Bulletin of Botanical Research, 2023, 43(4): 572-581.
[10]	Mingyue YUAN, Tianzhong ZHOU, Ma YU, Bin HU, Xiangyu LONG, Hua CHEN. Cloning and Functional Analysis of Deubiquitinating Enzyme Gene UCHs from Hevea brasiliensis [J]. Bulletin of Botanical Research, 2023, 43(4): 622-630.
[11]	Hong YANG, Lifeng WANG, Longjun DAI, Bingbing GUO. Effects of Tapping Panel Dryness on Mitochondrial Ultrastructure and ROS Metabolism in Barks of Rubber Tree (Hevea brasiliensis) [J]. Bulletin of Botanical Research, 2023, 43(1): 69-75.
[12]	Mengshuo LI, Yingze LIU, Huan LU, Sheng QIANG. Photosynthetic Capacity Differentiation and Gene Transcription in Different Geographical Populations of Arabidopsis thaliana under Common Garden conditions [J]. Bulletin of Botanical Research, 2023, 43(1): 90-99.
[13]	Bi QIN, Mingyang LIU, Meng WANG, Lifeng WANG, Fei HUANG. Cloning and Expression Analysis of DELLA gene HbRGL1 from Hevea brasiliensis Müll. Arg [J]. Bulletin of Botanical Research, 2022, 42(6): 997-1004.
[14]	Longjun DAI, Mingyang LIU, Jianghua YANG, Kai ZHOU, Bingbing GUO, Hong YANG, Lifeng WANG. Structural and Gene Expression Analysis of a DUF1262 Domain Protein in Latex from Hevea brasiliensis [J]. Bulletin of Botanical Research, 2022, 42(5): 802-810.
[15]	Sisi CHEN, Muhong XIE, Maokai CUI, Wenkai LI, Zhenggang XU, Caixia JIA, Guiyan YANG. Identification of Broussonetia papyrifera Transcription Factor BpbZIP1 and Analysis of Its Response to Cadmium Stress [J]. Bulletin of Botanical Research, 2022, 42(3): 394-402.