The Inositol-Phosphate Synthase Gene，MdMIPS1，Enhances Heat Stress Tolerance in Transgenic Apple Plants

doi:10.16420/j.issn.0513-353x.2024-0997

Acta Horticulturae Sinica ›› 2025, Vol. 52 ›› Issue (12): 3143-3156.doi: 10.16420/j.issn.0513-353x.2024-0997

• Genetic & Breeding·Germplasm Resources·Molecular Biology • Next Articles

The Inositol-Phosphate Synthase Gene，MdMIPS1，Enhances Heat Stress Tolerance in Transgenic Apple Plants

SUN Wenjing, LI Xiaoyan, ZHANG Jingyun, ZHOU Hui, HUANG Xiaojun, MA Fengwang^*(), GONG Xiaoqing^*()

State Key Laboratory for Crop Stress Resistance and High-Efficiency Production，Shaanxi Key Laboratory of Apple，College of Horticulture，Northwest A & F University，Yangling，Shaanxi 712100，China

Received:2025-08-08 Revised:2025-10-20 Online:2025-12-25 Published:2025-12-20
Contact: MA Fengwang, GONG Xiaoqing

Abstract

Abstract:

In order to reveal the role of Myo-inositol（MI）in apple plants responding to high-temperature stress，apple plants were first exogenously supplied with MI and treated with high temperature stress. The results showed that high-temperature stress significantly caused leaf yellow and wilting，increased the contents of reactive oxygen species（ROS），malondialdehyde（MDA），and relative electrolyte leakage（REL），decreased the contents of the chlorophyll in apple plants. However，100 µmol · L^-1exogenous MI treatment significantly alleviated the damage caused by high temperature. In addition，MdMIPS1 encoding the key rate-limiting enzyme in the MI biosynthesis，expressed highly under high-temperature stress in apple plants. The transgenic lines overexpressed MdMIPS1（OE，high MI content）were obtained in our previous study. They had higher endogenous content of MI，compared with wild type（WT）plants. And these OE lines were also used in this work and treated with high temperature stress. Compared with WT plants under high temperature stress，the OE lines exhibited slighter chlorisis and wilting in leaves. The contents of REL were lower and the contents of the chlorophyll were higher in OE plants than in WT. In addition，the stomatal aperture were larger in OE leaves，and the photosynthetic efficiency were stronger in OE plants than in WT. Meanwhile，the OE plants possessed less ROS accumulation and higher antioxidant enzyme activity than WT. The expression levels of several genes encoding the heat shock protein were also higher in OE plants than in WT under high temperature stress. In sum，all these results indicated that increasing the MI levels can significantly enhance tolerance to high temperature stress in apple plants.

Key words: apple, Myo-inositol, MdMIPS1, high-temperature stress

SUN Wenjing, LI Xiaoyan, ZHANG Jingyun, ZHOU Hui, HUANG Xiaojun, MA Fengwang, GONG Xiaoqing. The Inositol-Phosphate Synthase Gene，MdMIPS1，Enhances Heat Stress Tolerance in Transgenic Apple Plants[J]. Acta Horticulturae Sinica, 2025, 52(12): 3143-3156.

Figures/Tables 8

Table 1 Primer sequences of qRT-PCR

引物名称Primer name	正向引物序列（5′-3′）Forward primer sequence	反向引物序列Reverse primer sequence
qEF-1α	ATTCAAGTATGCCTGGGTGC	CAGTCAGCCTGTGATGTTCC
qMdMIPS1	AGGCCAAGTTCCACTCATTCCAC	CAAGCCCTCAGTATGTTCTCCAG
qMdHY5	AACTAGCATCCAAAGCTAGCC	CCACAGTCTGCAGTTTCACAAG
qMdDREB2A	CAGAGAACTTCACAACCGAC	CCAGTGCTCATGGCAAGGTTTT
qMdHSP17.3	AAGTTTGTGTTGCCGGAG	ATGTTATAATCCCTGCACGA
qMdHSP70-2	CTCTGTCTCTCGGTTTGGAG	GTCTTGTCCTCGGCAGAAA
qMdHSP70-1	CAAGCTGAAAGAGCTCAAGG	CTGTCCCAGCTGCATTACTT
qMdHSP90-5	ACACTCACTTCACCACTGAG	CATTTGAATCCACCACACCC

Fig. 1 Effect of exogenous MI on heat resistance of apple under high temperature stress A：The performance；B：MDA content；C：Total chlorophyll content；D：Electrolyte leakage. The data were shown as the mean of three replicates，and univariate analysis of variance and duncan multiple comparison were used，with different letters indicating significant differences between the data（P < 0.05）. The same below

Fig. 2 Effect of exogenous MI on antioxidant capacity of apple under high temperature stress

Fig. 3 Analysis of expression of apple MdMIPS1 A：Relative expression levels in tissues；B：Spatio-temporal expression patterns in leaves

Fig. 4 Analysis of heat resistance of MdMIPS1 transgenic apple plants under high temperature stress A：The performance of plant and leaf；B：Electrolyte leakage；C：Chlorophyll content. WT：GL-3 wild-type. OE-4/OE-6：Transgenic apple plants with overexpression of MdMIPS1. The same below

Fig. 5 Analysis of photosynthetic capacity of MdMIPS1 transgenic apple plants under high temperature stress A：The chlorophyll fluorescence；B：Fv/Fm；C：Tr；D：Pn；E：Ci；F：Leaf stomata；G：Stomatal opening；H：Gs. Different letters indicated WT，OE-4 and OE-6 strains have significant differences in control or heat stress data significant differences between the data（P < 0.05）

Fig. 6 Analysis of antioxidant capacity of MdMIPS1 transgenic apple plants under high temperature stress A：NBT and DAB staining；B： $\mathrm{O}_{2}^{\overline{·}}$ content；C：H2O2 content；D：SOD activity；E：POD activity

Fig. 7 Relative expression levels of heat shock proteins genes in WT and OE plants under high temperature stress Row Z-score normalization

References 45

[1]	Chen Yan, Han Liyuan, Du Zilong, Zhu Dongxing, Zhu Weimin, Zhang Yinging. 2024. Regulation mechanism of exogenous inositol improving high temperature stress tolerance of tomato seedlings. Acta Botanica Boreali Occidentalia Sinica, 44 (12):1856-1867. (in Chinese)
	陈艳, 韩利源, 杜子龙, 朱栋幸, 朱为民, 张迎迎. 2024. 外源肌醇提高番茄幼苗耐高温胁迫的调控机制. 西北植物学报, 44 (12):1856-1867.
[2]	Choudhury F K, Rivero R M, Blumwald E, Mittler R. 2017. Reactive oxygen species,abiotic stress and stress combination. Plant Journal, 90 (5):856-867.
[3]	Cochard H, Venisse J S, Barigah T S, Brunel N, Herbette S, Guilliot A, Tyree M T, Sakr S. 2007. Putative role of aquaporins in variable hydraulic conductance of leaves in response to light. Plant Physiol, 143 (1):122-133.
[4]	Cui Tengfei, Li Ting, Qu Mingshan. 2024. Effects of exogenous inositol on cold tolerance in mini watermelon. Chinese Agricultural Science Bulletin, 40 (25):66-71. (in Chinese)
	崔腾飞, 李婷, 曲明山. 2024. 外源肌醇对小型西瓜耐寒性的影响. 中国农学通报, 40 (25):66-71.
[5]	Cui Xiaoxia, Shi Chenle, Liu Xueqin. 2024. Effects of exogenous methyl jasmonate on seed germination and seedling physiology of soybean under cadmium stress. Chinese Journal of Soil and Fertilizer,(12):204-210. (in Chinese)
	崔晓霞, 石晨乐, 刘学勤. 2024. 外源茉莉酸甲酯对镉胁迫下大豆种子萌发及幼苗生理的影响. 中国土壤与肥料,(12):204-210.
[6]	Dai H Y, Li W R, Han G F, Yang Y, Ma Y, Li H, Zhang Z Z. 2013. Development of a seedling clone with high regeneration capacity and susceptibility to Agrobacterium in apple. Scientia Horticulturae, 164:202-208.
[7]	Das K, Roychoudhury A. 2014. Reactive oxygen species(ROS)and response of antioxidants as ROS-scavengers during environmental stress in plants. Frontiers in Environmental Science, 2:53.
[8]	de Pinto M C, Locato V, Paradiso A, de Gara L. 2015. Role of redox homeostasis in thermo-tolerance under a climate change scenario. Annals of Botany, 116:487-496
[9]	Degen G E, Orr D J, CarmoSilva E. 2020. Heat-induced changes in the abundance of wheat Rubisco activase isoforms. The New Phytologist, 229 (3):1298-1311.
[10]	Dhanda S S, Munjal R. 2012. Heat tolerance in relation to acquired thermotolerance for membrane lipids in bread wheat. Field Crops Res, 135:30-37.
[11]	Driedonks N, Xu J M, Peters J L, Park S, Rieu I. 2015. Multi-level interactions between heat shock factors,heat shock proteins,and the redox system regulate acclimation to heat. Frontiers in Plant Science, 6:999.
[12]	Fan X L, Zhang Z S, Gao H Y, Yang C, Liu M J, Li Y T, Li P M. 2014. Photoinhibition-like damage to the photosynthetic apparatus in plant leaves induced by submergence treatment in the dark. PLoS ONE, 9 (2):e89067.
[13]	Feder M E, Hofmann G E. 1999. Heat-shock proteins,molecular chaperones,and the stress response:evolutionary and ecological physiology. Annu Rev Physiol, 61:243-282.
[14]	Guo Yuanfei, Gan Lijun, Zhu Changhua, Li Guijun, Xia Kai. 2014. Effects of inositol on cold resistance of rice seedlings. Jiangsu Journal of Agricultural Sciences, 30 (6):1216-1221. (in Chinese)
	郭元飞, 甘立军, 朱昌华, 李桂俊, 夏凯. 2014. 肌醇对水稻幼苗抗寒性的影响. 江苏农业学报, 30 (6):1216-1221.
[15]	Hasanuzzaman M, Nahar K, Alam M M, Roychowdhury R, Fujita M. 2013. Physiological,biochemical,and molecular mechanisms of heat stress tolerance in plants. Int J Mol Sci, 14:9643-9684.
[16]	Hedhly A, Hormaza J, Herrero M. 2009. Global warming and sexual plant reproduction. Trends Plant Sci, 14:30-36.
[17]	Hu L Y, Zhou K, Ren G J, Yang S L, Liu Y, Zhang Z J, Li Y T S, Gong X Q, Ma F W. 2020b. Myo-inositol mediates reactive oxygen species-induced programmed cell death via salicylic acid-dependent and ethylene-dependent pathways in apple. Hortic Res, 7:138.
[18]	Hu S, Ding Y, Zhu C. 2020a. Sensitivity and responses of chloroplasts to heat stress in plants. Front Plant Sci, 11:375.
[19]	Huang H, Niu Y J, Cao H W, Tang X J, Xia X J, Yin W L, Dai S L. 2012. cDNA-AFLP analysis of salt inducible genes expression in Chrysanthemum lavandulifolium under salt treatment. Journal of Plant Physiology, 169 (4):410-414.
[20]	Ismail A, Takeda S, Nick P. 2014. Life and death under salt stress:same players,different timing? Journal of Experimental Botany, 65:2963-2979.
[21]	Jin J, Yang L, Fan D Y. 2019. Research progress on high temperature tolerance of fruit trees. Molecular Plant Breeding, 17 (3):1019-1027.
[22]	King A M, MacRae T H. 2015. Insect heat shock proteins during stress and diapause. Annu Rev Entomol, 60:59-75.
[23]	Klages K, Boldingh H, Smith G. 1999. Accumulation of myo-inositol in Actinidia seedlings subjected to salt stress. Annals of Botany, 84:521-527.
[24]	Li B J, Gao K, Ren H M, Tang W Q. 2018. Molecular mechanisms governing plant responses to high temperatures. Journal of Integrative Plant Biology, 60:757-779.
[25]	Li Jingqi, Li Xinlei. 2024. Effects of exogenous melatonin on seed germination of sunflower under NaCl stress. Shaanxi Journal of Agricultural Sciences, 70 (3):42-47. (in Chinese)
	李璟琦, 李欣蕾. 2024. 外源褪黑素对NaCl胁迫下向日葵种子萌发的影响. 陕西农业科学, 70 (3):42-47.
[26]	Lincoln Taiz, Eduardo Zeiger. 2015. Plant Physiology. Fifth Edition. 宋纯鹏,王学路,周云译. 北京: 科学出版社.
[27]	Liu J P, Zhang C C, Wei C C, Liu X, Wang M G, Yu F F, Xie Q, Tu J M. 2016. The RING finger ubiquitin E 3 ligase OsHTAS enhances heat tolerance by promoting H₂O₂-induced stomatal closure in rice. Plant Physiol, 170(1):429-443.
[28]	Luo Chunyan, Geng Hongkai, Wang Xiujun, et al. 2024. Effects of exogenous salicylic acid on the growth and photosynthesis of Ginkgo biloba under NaCl stress. Journal of Nanjing Forestry University(Natural Sciences Edition), 48 (6):91-101. (in Chinese)
	罗春燕, 耿红凯, 王秀军. 2024. NaCl胁迫下外源水杨酸对银杏生长和光合作用的影响. 南京林业大学学报(自然科学版), 48 (6):91-101.
[29]	Loewus F A, Murthy P P N. 2000. Myo-inositol metabolism in plants. Plant Science, 150:1-19.
[30]	Medina E, Kim S H, Yun M, Choi W G. 2021. Recapitulation of the function and role of ROS generated in response to heat stress in plants. Plants, 10:371.
[31]	Miao Tiantian, Li Qiang, Yu Hongjun, Liu Peng, Hao Jia, Jiang Weijie. 2021. Applied effect on cucumber seedling low temperature resistance of inositol. Journal of Chinese Vegetables,(2):72-79. (in Chinese)
	苗田田, 李强, 余宏军, 刘鹏, 郝佳, 蒋卫杰. 2021. 外施肌醇对黄瓜幼苗低温抗性的影响. 中国蔬菜,(2):72-79.
[32]	Ohama N, Sato H, Shinozaki K, Yamaguchi-Shinozaki K. 2017. Transcriptional regulatory network of plant heat stress response. Trends Plant Sci, 22:53-65.
[33]	Sihag P, Kumar U, Sagwal V, Kapoor P, Singh Y, Mehla S, Balyan P, Mir R R, Varshney R K, Singh K P, Dhankher O P. 2024. Effect of terminal heat stress on osmolyte accumulation and gene expression during grain filling in bread wheat(Triticum aestivum L.). The Plant Genome, 17 (1):e20307.
[34]	Teng Wei, Li Tingting, Ma Yan, Liu Xiyuan, Liu Ziming, Geng Wei. 2025. Alleviation effects of exogenous melatonin on melatonin seedlings under high-temperature stress and analysis of heat-tolerant gene expression. Journal of Northwest A & F University(Natural Science Edition),(7):1-8. (in Chinese)
	滕巍, 李婷婷, 马燕, 刘希元, 刘子铭, 耿伟. 2025. 外源褪黑素对高温胁迫甜瓜幼苗的缓解效应及耐热基因表达量分析. 西北农林科技大学学报(自然科学版),(7):1-8.
[35]	Wang X L, Xu C X, Cai X F, Wang Q H, Dai S J. 2017. Heat-responsive photosynthetic and signaling pathways in plants:insight from proteomics. International Journal of Molecular Sciences, 18 (10):2191.
[36]	Wei Hongyu, Song Zihui, He Shan, Liang Huiyuan, Yu Kunchi, Jian Wei, Peng Chunlin, Liu Yan, Han Jiajia, Zhao Zhengwu. 2024. Alleviation effects of exogenous carnosine on the growth of‘Shidao 960’rice seedlings under drought stress. Plant Physiology Journal, 60 (8):1280-1290. (in Chinese)
	魏红宇, 宋子慧, 何珊, 梁慧苑, 余昆驰, 简伟, 彭春霖, 刘艳, 韩佳佳, 赵正武. 2024. 外源肌肽对干旱胁迫‘师稻960’水稻幼苗生长的缓解效应. 植物生理学报, 60 (8):1280-1290.
[37]	Wei Xin, Gao Chunyan, Chang Chunhao, Tang Zhonghua. 2021. Metabolic regulation characteristics of exogenous inositol alleviating low temperature damage in Catharanthus roseus. Plant Physiology Journal, 57 (12):2247-2257. (in Chinese)
	魏鑫, 高春燕, 常纯皓, 唐中华. 2021. 外源肌醇缓解长春花低温伤害的代谢调控特征. 植物生理学报, 57 (12):2247-2257.
[38]	Xu Ying, Chen Jianhua, Zhu Aiguo, Ruan Mingbao, Wang Xiaofei, Sun Zhimin. 2015. Research progress of plant response mechanism under low temperature stress. Chinese Hemp Science, 37 (1):40-49. (in Chinese)
	许英, 陈建华, 朱爱国, 栾明宝, 王晓飞, 孙志民. 2015. 低温胁迫下植物响应机理的研究进展. 中国麻业科学, 37 (1):40-49.
[39]	Yang C, Zhang Z S, Gao H Y, Fan X L, Liu M J, Li X D. 2014. The mechanism by which NaCl treatment alleviates PSI photoinhibition under chilling-light treatment. J Photochem Photobiol B, 140:286-291.
[40]	Ye Y, Jia X, Xue M Y, Gao Y C, Yue H, Ma F W, Gong X Q. 2022. MpSnRK2.10 confers salt stress tolerance in apple via the ABA signaling pathway. Scientia Horticulturae, 298.
[41]	Yildizli A, Ünyayar S Ç S. 2018. Effects of exogenous myo-inositol on leaf water status and oxidative stress of Capsicum annuum under drought stress. Acta Physiologiae Plantarum, 40:122.
[42]	Yoshida T, Ohama N, Nakajima J, Kidokoro S, Mizoi J, Nakashima K, Maruyama K, Kim J M, Seki M, Todaka D, Osakabe Y, Sakuma Y, Schöffl F, Shinozaki K, Yamaguchi-Shinozaki K. 2011. Arabidopsis HsfA1 transcription factors function as the main positive regulators in heat shock-responsive gene expression. Mol Genet Genomics, 286:321-332.
[43]	You J, Chan Z L. 2015. ROS regulation during abiotic stress responses in crop plants. Frontiers in Plant Science, 6:109.
[44]	Zhang J, Yang N, Li Y, Zhu S, Zhang S, Sun Y, Zhang H X, Wang L, Su H. 2018. Overexpression of PeMIPS1 confers tolerance to salt and copper stresses by scavenging reactive oxygen species in transgenic poplar. Tree Physiology, 38 (10):1566-1577.
[45]	Zhang X L, Gong X Q, Su X J, Yu H X, Cheng S Y, Huang J W, Li D Y, Lei Z L, Li M J, Ma F W. 2023. The ubiquitin-binding protein MdRAD23D1 mediates drought response by regulating degradation of the proline-rich protein MdPRP6 in apple(Malus domestica). Plant Biotechnol J, 21:1560-1576.

The Inositol-Phosphate Synthase Gene，MdMIPS1，Enhances Heat Stress Tolerance in Transgenic Apple Plants

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