Functional Analysis of Soybean GmPM31 Gene Promoter Involvement in Response to High Temperature and Humidity Stress

doi:10.13560/j.cnki.biotech.bull.1985.2024-0435

Abstract

Abstract:

Objective To study the function of GmPM31 promoter on seed vigor formation in response to high temperature and high humidity stress, and to lay a foundation for comprehensively unravelling the function of soybean small heat shock proteins involved in the response to high temperature and high humidity stress. Method Homozygous T3 transgenic Arabidopsis thaliana with GmPM31 promoter was treated with high temperature and high humidity stress (40℃/100% relative humidity), and wild-type A. thaliana was used as control to detect its tolerance and stomatal opening. Analysis of GUS gene expression driven by the soybean GmPM31 promoter by GUS histochemical staining and quantitative real-time PCR. The seed vigor was detected by 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. Result Compared with the control group, the tolerance of GmPM31-transgenicpromotor A. thaliana to high temperature and high humidity increased. The activity of GUS increased, and they were expressed in the leaves, roots and flowers, and germination rate and vigor increased. Conclusion GmPM31 promoter improve the resistance of seeds to high temperature and high humidity stress and the ability to resist deterioration.

Key words: soybean, promoter, high temperature and high humidity, seed vigor

LIN Zi-yi, WU Yi-zhou, YE Fang-xian, ZHU Shu-ying, LIU Yan-min, LIU Su-shuang. Functional Analysis of Soybean GmPM31 Gene Promoter Involvement in Response to High Temperature and Humidity Stress[J]. Biotechnology Bulletin, 2025, 41(3): 90-97.

Figures/Tables 5

Fig. 1 Phenotypes of GmPM31-transgenic Arabidopsis plants under HTH stress

Fig. 2 Stomata assay of GmPM3-transgenic Arabidopsis plants under HTH conditionA: Stomatal morphology of WT and transgenic Arabidopsis grown for 28 d under HTH stress. B: Stomatal opening of WT and transgenic Arabidopsis under the HTH stress for 1 d and 2 d. C: Stomatal length of WT and transgenic Arabidopsis under the HTH stress for 1 d and 2 d. ** indicate that the differences are markly significant at 0.01 level. The same below

Fig. 3 Expression patterns of GUS genes driven by soybean GmPM31 gene promoter in various tissues and organs of ArabidopsisA: The control group of Arabidopsis grown for 25 d; B: the control group of Arabidopsis grown for 28 d; C: the control group of Arabidopsis grown for 25 d;D-H: the experimental group of Arabidopsis's flowers, blade, seeds, pod and root grown for 28 d, respectively; I: relative expressions of soybean GmPM31 gene promoter driving GUS genes in various tissues and organs of Arabidopsis. Different letters indicate the difference significant (P<0.01)

Fig. 4 Expression pattern of GUS genes driven by soybean GmPM31 gene promoter in Arabidopsis seedlingsA: GUS staining of Arabidopsis seedlings with GmPM31 promoter under normal condition; B: GUS staining of transgenic Arabidopsis seedlings with GmPM31 promoter treated with high temperature and humidity; C: relative expression of GUS genes driven by soybean GmPM31 gene promoter in Arabidopsis seedlings

Fig. 5 Effect of HTH stress on seed germination of GmPM31 transgenic ArabidopsisA: WT and GmPM31-transgenic Arabidopsis seedlings harvested after 7 d of sowing and then 2 d of HTH stress treatment; B: germination percentage of WT and GmPM3-transgenic Arabidopsis seeds under control conditions; C: germination percentage of WT and GmPM31-transgenic Arabidopsis seeds without HTH treatment; D: germination percentage of WT and GmPM31-transgenic Arabidopsis seeds after HTH treatment; E: TTC staining results of seeds after control and treatment groups

References 24

1	王文月, 姚志鹏, 于洋, 等. 我国大豆种业科技创新发展现状及对策建议 [J]. 中国农业科技导报, 2024, 26(3): 1-6.
	Wang WY, Yao ZP, Yu Y, et al. Scientific and technological innovation of soybean seed industry in China: current situation and strategy [J]. J Agric Sci Technol, 2024, 26(3): 1-6.
2	舒英杰, 周玉丽, 陶源, 等. 生理成熟期高温高湿胁迫对春大豆种子生理特性及种子劣变相关基因GmSbh1表达的影响 [J]. 生态学杂志, 2016, 35(12): 3286-3292.
	Shu YJ, Zhou YL, Tao Y, et al. Effects of high temperature and humidity stress on the physiological characters of spring soybean seeds and the expression of seed deterioration-related gene GmSbh1 at the physiological maturity stage [J]. Chin J Ecol, 2016, 35(12): 3286-3292.
3	邢悦楠, 王皓, 张琼, 等. 植物小分子热激蛋白的功能及表达调控 [J]. 植物生理学报, 2023, 59(8): 1524-1532.
	Xing YN, Wang H, Zhang Q, et al. Function and expression regulation of plant small molecule heat shock proteins [J]. Plant Physiol J, 2023, 59(8): 1524-1532.
4	刘骕骦, 邱颖胜, 刘燕敏, 等. 大豆GmPM31基因生物信息学、组织表达及高温高湿响应分析 [J]. 大豆科学, 2021, 40(5): 612-619.
	Liu SS, Qiu YS, Liu YM, et al. Analysis of soybean GmPM31 bioinformatics, tissue expression and response to high temperature and high humidity [J]. Soybean Sci, 2021, 40(5): 612-619.
5	刘骕骦, 李昕雨, 王泽博, 等. 大豆GmPM31基因的功能分析及拟南芥转化 [J]. 大豆科学, 2022, 41(5): 526-535.
	Liu SS, Li XY, Wang ZB, et al. Functional analysis of soybean GmPM31 gene and transformation into Arabidopsis thaliana [J]. Soybean Sci, 2022, 41(5): 526-535.
6	陈建琦, 赵明珠, 王义, 等. 植物中组织特异性启动子的研究进展 [J]. 北方园艺, 2023(19): 128-134.
	Chen JQ, Zhao MZ, Wang Y, et al. Research progress on tissue specific promoters in plant [J]. North Hortic, 2023(19): 128-134.
7	Zhou ZX, Wang J, Yu QH, et al. Promoter activity and transcriptome analyses decipher functions of CgbHLH001 gene (Chenopodium glaucum L.) in response to abiotic stress [J]. BMC Plant Biol, 2023, 23(1): 116.
8	Dong GQ, Fan MW, Wang HN, et al. Functional characterization of TkSRPP promoter in response to hormones and wounding stress in transgenic tobacco [J]. Plants (Basel), 2023, 12(2): 252.
9	Huang JJ, Yan XT, Li JJ, et al. OsRhoGAP2 promoter drives inflorescence-preferential expression and confers responses to abiotic stresses in transgenic Arabidopsis [J]. Acta Physiol Plant, 2019, 41(5): 67.
10	刘骕骦. 大豆基质金属蛋白酶基因Gm1-MMP和Gm2-MMP的分离以及响应高温高湿胁迫的功能分析 [D]. 南京: 南京农业大学, 2017.
	Liu SS. Isolation of soybean matrix metalloproteinase genes Gm1-MMP and Gm2-MMP and functional analysis in response to high temperature and high humidity stress [D]. Nanjing: Nanjing Agricultural University, 2017.
11	Waters ER, Vierling E. Plant small heat shock proteins - evolutionary and functional diversity [J]. New Phytol, 2020, 227(1): 24-37.
12	Neto VG, Barbosa RR, Carosio MGA, et al. Sequence analysis of Ricinus communis small heat-shock protein (sHSP) subfamily and its role in abiotic stress responses [J]. Ind Crops Prod, 2020, 152: 112541.
13	Lemonnier P, Lawson T. Calvin cycle and guard cell metabolism impact stomatal function [J]. Semin Cell Dev Biol, 2024, 155: 59-70.
14	Ma W, Li J, Liu FJ, et al. GhHSP24.7 mediates mitochondrial protein acetylation to regulate stomatal conductance in response to abiotic stress in cotton [J]. Crop J, 2023, 11(4): 1128-1139.
15	Zhong LS, Shi YX, Xu SL, et al. Heterologous overexpression of heat shock protein 20 genes of different species of yellow Camellia in Arabidopsis thaliana reveals their roles in high calcium resistance [J]. BMC Plant Biol, 2024, 24(1): 5.
16	Zhang YL, Sun YL, Liu XJ, et al. Populus euphratica apyrases increase drought tolerance by modulating stomatal aperture in Arabidopsis [J]. Int J Mol Sci, 2021, 22(18): 9892.
17	Kaundal A, Ramu VS, Oh S, et al. GENERAL CONTROL NONREPRESSIBLE4 degrades 14-3-3 and the RIN4 complex to regulate stomatal aperture with implications on nonhost disease resistance and drought tolerance [J]. Plant Cell, 2017, 29(9): 2233-2248.
18	Wu JT, Gao T, Hu JN, et al. Research advances in function and regulation mechanisms of plant small heat shock proteins (sHSPs) under environmental stresses [J]. Sci Total Environ, 2022, 825: 154054.
19	González-Gordo S, Palma JM, Corpas FJ. Small heat shock protein (sHSP) gene family from sweet pepper (Capsicum annuum L.) fruits: involvement in ripening and modulation by nitric oxide (NO) [J]. Plants (Basel), 2023, 12(2): 389.
20	Wang P, Zhang TT, Li YX, et al. Comprehensive analysis of Dendrobium catenatum HSP20 family genes and functional characterization of DcHSP20-12 in response to temperature stress [J]. Int J Biol Macromol, 2024, 258(Pt 2): 129001.
21	Ding XL, Lv ML, Liu Y, et al. A small heat shock protein GmHSP18.5a improves the male fertility restorability of cytoplasmic male sterility-based restorer line under high temperature stress in soybean [J]. Plant Sci, 2023, 337: 111867.
22	Guo LM, Li J, He J, et al. A class I cytosolic HSP20 of rice enhances heat and salt tolerance in different organisms [J]. Sci Rep, 2020, 10(1): 1383.
23	Mukesh Sankar S, Tara Satyavathi C, Barthakur S, et al. Differential modulation of heat-Inducible genes across diverse genotypes and molecular cloning of a sHSP from pearl Millet[Pennisetum glaucum (L.) R. br.] [J]. Front Plant Sci, 2021, 12: 659893.
24	Ma W, Zhao T, Li J, et al. Identification and characterization of the GhHsp20 gene family in Gossypium hirsutum [J]. Sci Rep, 2016, 6: 32517.

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