Functional Analysis of StHY5 Associated with Low-Temperature Stress in Potato

doi:10.16420/j.issn.0513-353x.2024-0723

Acta Horticulturae Sinica ›› 2025, Vol. 52 ›› Issue (7): 1745-1757.doi: 10.16420/j.issn.0513-353x.2024-0723

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

Functional Analysis of StHY5 Associated with Low-Temperature Stress in Potato

DOU Xueting¹, ZHU Xi²^,³, ZHANG Ning¹^,^*(), and SI Huaijun¹

¹ State Key Laboratory of Aridland Crop Science，College of Life Science and Technology，Gansu Agricultural University，Lanzhou 730070，China
² Key Laboratory of Tropical Fruit Biology，Ministry of Agriculture and Rural Affairs/Hainan Provincial Key Laboratory of Tropical Horticulture Postharvest Treatment and Preservation，Institute of South Subtropical Crops，Chinese Academy of Tropical Agricultural Sciences，Zhanjiang，Guangdong 524091，China
³ National Key Laboratory of Biological Breeding of Tropical Crops，Sanya Research Institute，Chinese Academy of Tropical Agricultural Sciences，Sanya，Hainan 572024，China

Received:2024-11-12 Revised:2025-04-22 Online:2025-07-23 Published:2025-07-23
Contact: ZHANG Ning

Abstract

Abstract:

HY5 is a class of bZIP transcription factor. HY5 is involved in regulating plant light response and abiotic stress. In this study，the functions of the StHY5 and its encoded protein were analyzed，and the subcellular localization was observed using potato cultivar‘Atlantic’as the experimental material. The expression pattern of StHY5 gene was analyzed under 4 ℃ low-temperature stress by qRT-PCR method. The overexpression and interference expression vectors of StHY5 gene were constructed，and obtained transgenic potato plants through genetic transformation technology. The changes of reactive oxygen species under 4 ℃ low-temperature treatment. The results showed that the coding sequence of StHY5 gene was 477 bp，encoding 158 amino acids. StHY5 protein was located in the nucleus，and it belonged to unstable hydrophilic proteins，without signal peptide and transmembrane structure. It had conserved evolutionary among different species，and had the highest evolutionary relationship with wild tomato and Solanum stenotomum. StHY5 protein was closely related to 10 proteins interaction，including 1 photosensitive pigment A，2 B-box proteins，and 3 histone deacetylases. There were cis-acting elements involved in light-responsive and hormone-responsive in the promoter region of StHY5 gene. The qRT-PCR results showed that StHY5 gene was consistently up-regulated in expression during low-temperature. The superoxide dismutase（SOD），peroxidase（POD），catalase（CAT）activities and proline（Pro）content of StHY5 gene overexpression plants were significantly higher and the malondialdehyde（MDA）content was significantly lower than wild type plants under low-temperature treatment. However，StHY5 gene interference expression plants was reversed under low-temperature treatment.

Key words: potato, StHY5, low temperature, bioinformatics analysis, genetic transformation

DOU Xueting, ZHU Xi, ZHANG Ning, and SI Huaijun. Functional Analysis of StHY5 Associated with Low-Temperature Stress in Potato[J]. Acta Horticulturae Sinica, 2025, 52(7): 1745-1757.

Figures/Tables 11

Table 1 Specific primers for PCR and qRT-PCR in this study

用途Usage	引物名称Primer name	引物序列Primer sequence（5′-3′）
基因克隆 Gene cloning	StHY5-F	CGGGGGACGAGCTCGGTACCATGCAAGAGCAAGCGACAAGTTC
	StHY5-R	TGCTCACCATGTCGACTTACTTCCTCCCTTCCTTTGCACC
	pMD18-StHY5-F	CGGGGGACGAGCTCGGTACCGGGCCCCCCCTCGAGGTCG
	pMD18-StHY5-R	CCATGTCGACTCTAGAACTAGTGGATCCCCCCATGGC
	StHY5-YF	CGGGGGACGAGCTCGGTACCATGCAAGAGCAAGCGACAAGTTC
	StHY5-YR	TGCTCACCATGTCGACCTTCCTCCCTTCCTTTGCACC
	HYG-F	GCTTCTGCGGGCGATTTGTGT
	HYG-R	GGTCGCGGAGGCTATGGATGC
实时荧光定量PCR qRT-PCR	StHY5-qF	CAGCAAGCAAGGGAGAGGAA
实时荧光定量PCR qRT-PCR	StHY5-qR	TTCCTCCCTTCCTTTGCACC
内参基因 Reference gene	StEf1α-F	GATGGTCAGACCCGTGAACA
内参基因 Reference gene	StEf1α-R	CCTTGGAGTACTTCGGGGTG

Table 2 Primers of amiR-StHY5 for PCR

引物名称Primer name	引物序列（5′-3′）Primer sequence
Ⅰ	gaTTTCAGTATATGTCTAAGCGTtctctcttttgtattcc
Ⅱ	gaACGCTTAGACATATACTGAAAtcaaagagaatcaatga
Ⅲ	gaACACTTAGACATAAACTGAATtcacaggtcgtgatatg
Ⅳ	gaATTCAGTTTATGTCTAAGTGTtctacatatatattcct
A	CTGCAAGGCGATTAAGTTGGGTAAC
B	GCGGATAACAATTTCACACAGGAAACAG

Fig. 1 Secondary structure（A）and tertiary structure（B）of StHY5 protein

Fig. 2 Multi-sequence alignment of StHY5 protein and other homologous proteins of species

Fig. 3 StHY5 protein phylogenetic evolutionary tree

Fig. 4 StHY5 gene amplification product band（A）and amiR-StHY5 precursor fragment（B）

Fig. 5 Subcellular localization of pCEGFP-StHY5

Fig. 6 Relative expression of StHY5 gene under low temperature treatment The error line indicates the standard error，different lowercase letters represent significant differences（P < 0.05）

Fig. 7 PCR identification of transgenic plants +：Positive control；-：Negative control

Fig. 8 Relative expression of the transgenic plants by qRT-PCR WT：Wild type. The error line indicates the standard error，different lowercase letters represent significant differences（P < 0.05），the same below

Fig. 9 Physiological indexes of the transgenic plants under low temperature stress

References 40

[1]	Ang L H, Chattopadhyay S, Wei N, Oyama T, Okada K, Batschauer A, Deng X W. 1998. Molecular interaction between COP1 and HY 5 defines a regulatory switch for light control of Arabidopsis development. Molecular Cell, 1 (2):213-222. doi: 10.1016/s1097-2765(00)80022-2 pmid: 9659918
[2]	Boyer J S. 1982. Plant productivity and environment. Science, 218 (4571):443-448. doi: 10.1126/science.218.4571.443 pmid: 17808529
[3]	Burman N, Bhatnagar A, Khurana J P. 2018. OsbZIP48,a HY 5 transcription factor ortholog,exerts pleiotropic effects in light-regulated development. Plant Physiology, 176 (2):1262-1285. doi: 10.1104/pp.17.00478 pmid: 28775143
[4]	Catalá R, Medina J, Salinas J. 2011. Integration of low temperature and light signaling during cold acclimation response in Arabidopsis. Proceedings of the National Academy of Sciences, 108 (39):16475-16480.
[5]	Chen X, Yao Q, Gao X, Jiang C, Harberd N P, Fu X. 2016. Shoot-to-root mobile transcription factor HY 5 coordinates plant carbon and nitrogen acquisition. Current Biology, 26 (5):640-646.
[6]	Cluis C P, Mouchel C F, Hardtke C S. 2004. The Arabidopsis transcription factor HY 5 integrates light and hormone signaling pathways. The Plant Journal, 38 (2):332-347.
[7]	Deng Fuyuan, Qiao Zhongquan, Wang Xiaoming, Li Chunxia, Lu Liushu, Li Xuelu, He Gang. 2022. Cloning,subcellular localization and expression analysis of LiHY5 gene in Lagerstroemia indica under different light quality. Journal of Jiangxi Agricultural University, 44 (6):1546-1554. (in Chinese)
	邓涪元, 乔中全, 王晓明, 李春霞, 陆柳淑, 李雪露, 何钢. 2022. 紫薇LiHY5基因的克隆、亚细胞定位与不同光质下的表达分析. 江西农业大学学报, 44 (6):1546-1554.
[8]	Deng Shufang, Liu Qian, Liu Ling, Chen Ou, Wang Wenjun, Zeng Kaifang, Deng Lili. 2024. Cloning of mandarin fruit CcHY5 and its function in fruit coloration. Acta Horticulturae Sinica, 51 (5):939-955. (in Chinese)
	邓淑芳, 刘倩, 刘玲, 陈鸥, 王文军, 曾凯芳, 邓丽莉. 2024. 蜜橘CcHY5的克隆及其对果实转色功能的研究. 园艺学报, 51 (5):939-955.
[9]	Deng Yingyi, Zheng Xu, Xiong Jun, Xu Juan, Qin Weizhi. 2017. Identification of cold resistance of new potato variety Guinongshu No.1 winter seed in the field. Southern Journal of Agriculture, 48 (1):66-71. (in Chinese)
	邓英毅, 郑虚, 熊军, 许娟, 覃维治. 2017. 马铃薯新品种桂农薯1号冬种田间耐寒性鉴定. 南方农业学报, 48 (1):66-71.
[10]	Foley R C, Grossman C, Ellis J G, Llewellyn D J, Dennis E S, Peacock W J, Singh K B. 1993. Isolation of a maize bZIP protein subfamily:candidates for the ocs-element transcription factor. The Plant Journal, 3 (5):669-679.
[11]	Franklin K A, Toledo-Ortiz G, Pyott D E, Halliday K J. 2014. Interaction of light and temperature signaling. Journal of Experimental Botany, 65 (11):2859-2871. doi: 10.1093/jxb/eru059 pmid: 24569036
[12]	Gangappa S N, Botto J F. 2016. The multifaceted roles of HY 5 in plant growth and development. Molecular Plant, 9 (10):1353-1365.
[13]	Kim S, Hwang G, Lee S, Zhu JY, Paik I, Nguyen T T, Kim J, Oh E. 2017. High ambient temperature represses anthocyanin biosynthesis through degradation of HY5. Frontiers in Plant Science,8:1787.
[14]	Koornneef M, Rolff E, Spruit C J P. 1980. Genetic control of light-inhibited hypocotyl elongation in Arabidopsis thaliana(L.) Heynh. Zeitschrift für Pflanzenphysiologie, 100 (2):147-160.
[15]	Li J, Li G, Gao S, Martinez C, He G, Zhou Z, Huang X, Lee J, Zhang H, Shen Y, Wang H, Deng X W. 2010. Arabidopsis transcription factor ELONGATED HYPOCOTYL5 plays a role in the feedback regulation of phytochrome A signaling. The Plant Cell, 22 (11):3634-3649.
[16]	Li Y, Shi Y, Li M, Fu D, Wu S, Li J, Gong Z, Liu H, Yang S. 2021. The CRY2-COP1-HY5-BBX7/ 8 module regulates blue light-dependent cold acclimation in Arabidopsis. The Plant Cell, 33 (11):3555-3573.
[17]	Liu W T, Zhang L C, Ma L, Yuan L, Wang A D. 2023. The HY 5 transcription factor negatively regulates ethylene production by inhibiting ACS1 expression under blue light conditions in pear. Horticultural Plant Journal, 9 (5):920-930.
[18]	Liu X, Wei J, Li S, Li J, Cao H, Huang D, Zhang D, Zhang Z, Gao T, Zhang Y, Ma F, Li C. 2024. MdHY 5 positively regulates cold tolerance in apple by integrating the auxin and abscisic acid pathways. New Phytologist, 246 (5):2155-2173.
[19]	Liu Y, Roof S, Ye Z, Giovannoni J. 2004. Manipulation of light signal transduction as a means of modifying fruit nutritional quality in tomato. Proceedings of the National Academy of Sciences, 101 (26):9897-9902.
[20]	Ma R, Tian N, Wang J, Fan M, Wang B, Qu P, Xu S, Xu Y, Cheng C, Lv P. 2022a. Genome-wide identification and characterization of banana Ca²⁺-ATPase genes and expression analysis under different concentrations of Ca²⁺ treatments. International Journal of Molecular Sciences, 23 (19):11914.
[21]	Ma X, Gai WX, Li Y, Yu YN, Ali M, Gong ZH. 2022b. The CBL-interacting protein kinase CaCIPK13 positively regulates defence mechanisms against cold stress in pepper. Journal of Experimental Botany, 73 (5):1655-1667.
[22]	Nijhawan A, Jain M, Tyagi A K, Khurana J P. 2008. Genomic survey and gene expression analysis of the basic leucine zipper transcription factor family in rice. Plant Physiology, 146 (2):333-350. doi: 10.1104/pp.107.112821 pmid: 18065552
[23]	Nishimura R, Ohmori M, Fujita H, Kawaguchi M. 2002. A Lotus basic leucine zipper protein with a RING-finger motif negatively regulates the developmental program of nodulation. Proceedings of the National Academy of Sciences, 99 (23):15206-15210.
[24]	Oyama T, Shimura Y, Okada K. 1997. The Arabidopsis HY5 gene encodes a bZIP protein that regulates stimulus-induced development of root and hypocotyl l. Genes & Development, 11 (22):2983-2995.
[25]	Petersen H V, Serup P, Leonard J, Michelsen B K, Madsen O D. 1994. Transcriptional regulation of the human insulin gene is dependent on the homeodomain protein STF1/IPF1 acting through the CT boxes. Proceedings of the National Academy of Sciences, 91 (22):10465-10469.
[26]	Si Huaijun, Xie Conghua, Liu Jun. 2003. An efficient protocol for agrobacterium-mediated transformation with microtuber and the introduction of an antisense class I patatin gene into potato. Acta Agronomica Sinica, 29 (6):801-805. (in Chinese)
	司怀军, 谢从华, 柳俊. 2003. 农杆菌介导的马铃薯试管薯遗传转化体系的优化及反义class I patatin基因的导入. 作物学报, 29 (6):801-805.
[27]	Song J, Lin R, Tang M, Wang L, Fan P, Xia X, Yu J, Zhou Y. 2023. SlMPK1-and SlMPK2-mediated SlBBX17 phosphorylation positively regulates CBF-ependent cold tolerance in tomato. New Phytologist, 239 (5):1887-1902.
[28]	Tan Zhengwei, Lu Dandan, Li Lei, Yu Yongliang, Xu Lanjie, Yang Hongqi, Yang Qing, Dong Wei, Li Chunming, An Sufang, Lu Hailing, Liang Huizhen. 2022. Cloning and expression analysis of CtHY5,a key gene of light-regulated signaling pathway in safflower. Chinese Herbal Medicine, 53 (18):5825-5833. (in Chinese)
	谭政委, 鲁丹丹, 李磊, 余永亮, 许兰杰, 杨红旗, 杨青, 董薇, 李春明, 安素妨, 芦海灵, 梁慧珍. 2022. 红花光调控信号途径关键基因CtHY5的克隆及表达分析. 中草药, 53 (18):5825-5833.
[29]	Ulm R, Baumann A, Oravecz A, Nagy F. 2004. Genome-wide analysis of gene expression reveals function of the bZIP transcription factor HY5 in the UV-B response of Arabidopsis. Proceedings of the National Academy of Sciences, 101 (5):1397-1402.
[30]	Wang F, Zhang L, Chen X, Wu X, Xiang X, Zhou J, Xia X, Shi K, Yu J, Foyer C H. 2019. SlHY5integrates temperature,light,and hormone signaling to balance plant growth and cold tolerance. Plant Physiology, 179 (2):749-760.
[31]	Wang Mengyao, Wang Mengjie, Lai Huiping, He Yaping, Yan Lu, Li Peng, Guo Liting, Ai Ye. 2024. Research progress on the effect of temperature on anthocyanin accumulation in plants. Acta Horticulturae Sinica, 51 (7):1501-1515. (in Chinese) doi: 10.16420/j.issn.0513-353x.2023-0556
	王梦瑶, 王梦洁, 赖慧萍, 贺雅萍, 鄢璐, 黎鹏, 郭丽婷, 艾叶. 2024. 温度影响植物花青苷积累的研究进展. 园艺学报, 51 (7):1501-1515.
[32]	Wang Z, Cheng K, Wan L, Yan L, Jiang H, Liu S, Lei Y, Liao B. 2015. Genome-wide analysis of the basic leucine zipper(bZIP)transcription factor gene family in six legume genomes. BMC Genomics,16:1053.
[33]	Weller J L, Hecht V, Vander Schoor J K, Davidson S E, Ross J J. 2009. Light regulation of gibberellin biosynthesis in pea is mediated through the COP1/HY 5 pathway. The Plant Cell, 21 (3):800-813.
[34]	Xu D. 2020. COP1 and BBXs‐HY5‐mediated light signal transduction in plants. New Phytologist, 228 (6):1748-1753.
[35]	Yamawaki S, Yamashino T, Nakanishi H, Mizuno T. 2011. Functional characterization of HY 5 homolog genes involved in early light-signaling in Physcomitrella patens. Bioscience,Biotechnology,and Biochemistry, 75 (8):1533-1539.
[36]	Yang Ying, Gao Shiqing, Tang Yimiao, Ye Xiaofang, Wang Yongbo, Liu Meiying, Zhao Changping. 2009. Research progress on BZIP transcription factors in plants. Journal of Triticeae Crops, 29 (4):730-737. (in Chinese)
	杨颖, 高世庆, 唐益苗, 冶晓芳, 王永波, 刘美英, 赵昌平. 2009. 植物bZIP转录因子的研究进展. 麦类作物学报, 29 (4):730-737.
[37]	Zhang Chao, Zhang Hui, Zhao Xiaoyan, Ma Yue, Yao Huiyuan. 2009. Secondary structure of winter wheat bran antifreeze proteins using circular dichroism spectroscopy and infrared spectroscopy. Spectroscopy and Spectral Analysis, 29 (7):1764-1767. (in Chinese) pmid: 19798935
	张超, 张晖, 赵晓燕, 马越, 姚惠源. 2009. 使用圆二色性光谱和红外光谱研究冬小麦麸皮抗冻蛋白的二级结构. 光谱学与光谱分析, 29 (7):1764-1767. pmid: 19798935
[38]	Zhang L, Jiang X, Liu Q, Ahammed G J, Lin R, Wang L, Shao S, Yu J, Zhou Y. 2020. The HY5 and MYB 15 transcription factors positively regulate cold tolerance in tomato via the CBF pathway. Plant,Cell & Environment, 43 (11):2712-2726.
[39]	Zhang Li, Zhou Bo, Li Yuhua. 2010. Research progress on the structure and function of plant HY 5 protein. Plant Physiol Communications, 46 (10):985-990. (in Chinese)
	张荔, 周波, 李玉花. 2010. 植物HY5蛋白结构与功能的研究进展. 植物生理学通讯, 46 (10):985-990.
[40]	Zhang Y, Liu Z, Liu R, Hao H, Bi Y. 2011. Gibberellins negatively regulate low temperature-induced anthocyanin accumulation in a HY5/HYH-dependent manner. Plant Signaling & Behavior, 6 (5):632-634.

Functional Analysis of StHY5 Associated with Low-Temperature Stress in Potato

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