月季‘月月红’RcRAP2.7的克隆及其在非生物胁迫下的表达分析

doi:10.16420/j.issn.0513-353x.2024-0532

摘要/Abstract

摘要： 为探究‘月月红’（Rosa chinensis‘Yueyuehong’）月季中AP2/ERF转录因子家族成员RcRAP2.7对非生物胁迫的响应，克隆了RcRAP2.7 CDS全长，并进行转录表达分析。RcRAP2.7 CDS序列全长1 338 bp，编码446个氨基酸，在121 ~ 183和212 ~ 276位氨基酸区域含有两个AP2/ERF保守结构域。亚细胞定位结果显示RcRAP2.7蛋白定位于细胞膜和细胞核。启动子区域调控元件分析表明RcRAP2.7可能涉及激素信号通路和胁迫响应。进一步转录激活活性分析表明，其C端结构域具有转录激活活性。通过实时荧光定量PCR分析发现，RcRAP2.7在干旱、高盐和极端温度等胁迫条件下，其表达模式存在差异：在叶片中，RcRAP2.7在高温和干旱胁迫下较对照下调表达，而在低温和高盐胁迫下先上调后下调又上调；在根中，RcRAP2.7表达量受干旱、高盐和极端温度等胁迫的影响，均随着胁迫时间的增加呈现逐渐上升趋势，在胁迫48 h时表达量最高。

关键词: 月季, RcRAP2.7, 非生物胁迫, 表达模式

Abstract: In order to investigate the role of RcRAP2.7，a member of the AP2/ERF transcription factor family，in response to abiotic stress in Rosa chinensis‘Yueyuehong’. In this study，the full length of the coding sequence（CDS）of RcRAP2.7 was cloned，and then it was subjected to transcriptional expression analysis. The results showed that the CDS of RcRAP2.7 was 1 338 bp in length，encoding 446 amino acids，and there were two AP2/ERF conserved domains，located in 121–183 aa and 212–276 aa. And the RcRAP2.7 protein was located at the cell membrane and nucleus. Analysis of the promoter region regulatory elements indicated that RcRAP2.7 may be involved in hormone signalling pathways and stress response. Further transcriptional activation activity analysis showed that its C-terminal structural domain possessed transcriptional activation activity. Quantitative real-time PCR revealed that the expression pattern of RcRAP2.7 differed under various stress conditions such as drought，high salt and extreme temperature. In the leaves，the expression of RcRAP2.7 was down-regulated in high temperature and drought stress as a whole compared with the control，while it was first up-regulated，then down-regulated and then up-regulated in low temperature and high salt stress compared with the control. In the roots，its expression was affected by drought，high salt and extreme temperature，all of which showed a gradual increase with increasing treatment time，with the highest expression at 48 h of stress treatment.

Key words: Rosa chinensis, RcRAP2.7, abiotic stress, expression mode

中图分类号:

S 685.12

马豫皖, 刘澳, 刘向东, 张雅婧, 董炫克, 李玉帆, 陈己任. 月季‘月月红’RcRAP2.7的克隆及其在非生物胁迫下的表达分析[J]. 园艺学报, 2025, 52(4): 921-932.

MA Yuwan, LIU Ao, LIU Xiangdong, ZHANG Yajing, DONG Xuanke, LI Yufan, CHEN Jiren. Cloning of the RcRAP2.7 Gene and its Expression Analysis Under Abiotic Stresses in Rosa chinensis‘Yueyuehong’[J]. Acta Horticulturae Sinica, 2025, 52(4): 921-932.

参考文献

[1]	Ai Rong，Zhang Chun，Yue Manfang，Zou Huawen，Wu Zhongyi. 2023. Response of maize transcriptional factor ZmEREB211 to abiotic stress. Acta Agronomica Sinica，49 (9)：2433–2445. (in Chinese)
	艾蓉，张春，悦曼芳，邹华文，吴忠义. 2023. 玉米转录因子ZmEREB211对非生物逆境胁迫的应答. 作物学报，49 (9)：2433–2445.
[2]	Bian X F，Kim H S ，Kwak S S，Zhang Q，Liu S，Ma P Y，Jia Z D，Xie Y Z，Zhang P，Yu Y. 2022. Different functions of IbRAP2.4，a drought-responsive AP2/ERF aranscription factor，in regulating root development between Arabidopsis and sweetpotato. Frontiers in Plant Science，13：820450.
[3]	Cai X Y，Magwanga R O，Xu Y C，Zhou Z L，Wang X X，Hou Y Q，Wang Y H，Zhang Y M，Liu F，Wang K B. 2019. Comparative transcriptome，physiological and biochemical analyses reveal response mechanism mediated by CBF4 and ICE2 in enhancing cold stress tolerance in Gossypium thurberi. AoB PLANTS，11 (6)：plz045.
[4]	Chen J R，Chen Y B，Ziemiańska M，Liu R，Deng Z N，NiedźwieckaFilipiak I，Li Y L，Jiao J X，Xiong X Y. 2016. Co-expression of MtDREB1C and RcXET enhances stress tolerance of transgenic China rose（Rosa chinensis Jacq.）. Journal of Plant Growth Regulation，35 (2)：586–599.
[5]	Cui bo，Hao Ping’an，Liang Fang，Zhang Yan，Wang Ximeng，Li Junlin，Jiang Suhua，Xu Shenping. 2020. Cloning and expression analysis of AP2/ERF family gene from Phalaenopsis under low temperature. Acta Horticulturae Sinica，47 (1)：85–97. (in Chinese)
	崔波，郝平安，梁芳，张燕，王喜蒙，李俊霖，蒋素华，许申平. 2020. 蝴蝶兰AP2/ERF 家族基因的克隆及在低温下表达特性分析. 园艺学报，2020，47 (1)：85–97.
[6]	Dong C J，Liu J Y. 2010. The Arabidopsis EAR-motif-containing protein RAP2.1 functions as an active transcriptional repressor to keep stress responses under tight control. BMC Plant Biology，10：1–15.
[7]	Dong Cuicui，Ma Yanyan，Xie Rangjin，Deng Lie，Yi Shilai，Lü Qiang，Zheng Yongqiang，He Shaolan. 2016. Expression of two citrus AP2/ERF genes under different hormone and stress treatments. Acta Horticulturae Sinica，43 (2)：239–248. (in Chinese)
	董翠翠，马岩岩，谢让金，邓烈，易时来，吕强，郑永强，何绍兰. 2016. 柑橘CitERF9和CitAP2-7在不同逆境和外源激素处理下的表达. 园艺学报，43 (2)：239–248.
[8]	Geng L F，Yu S，Zhang Y C，Su L，Lu W P，Zhu H，Jiang X Q. 2023. Transcription factor RcNAC091 enhances rose drought tolerance through the abscisic acid-dependent pathway. Plant Physiology，193 (2)：1695–1712.
[9]	Hinz M，Wilson I W，Yang J，Buerstenbinder K，Llewellyn D，Dennis E S，Sauter M，Dolferus R. 2010. Arabidopsis RAP2.2：an ethylene response transcription factor that is important for hypoxia survival. Plant Physiology，153 (2)：757–772.
[10]	Jia Xin，Zeng Zhen，Chen Yue，Feng Hui，Lü Yingmin，Zhao Shiwei. 2022. Cloning and expression analysis of RcDREB2A gene in Rosa chinensis ‘Old Blush’. Acta Horticulturae Sinica，49 (9)：1945–1956. (in Chinese)
	贾鑫，曾臻，陈月，冯慧，吕英民，赵世伟. 2022. 月季‘月月粉’RcDREB2A的克隆与表达分析. 园艺学报，49 (9)：1945–1956.
[11]	Lan Mengjiao，Hou Meng，Xiao Manqiu，Li Chen，Pan Hao，Zhang Yungang，Lu Lingzhi，Hou Longying，Ge Ruihua，Wu Wensheng，Li Qiang. 2023. Research progress of AP2/ERF transcription factors participating in plant secondary metabolism and stress response. Journal of Plant Genetic Resources，24 (5)：1223–1235. (in Chinese)
	兰孟焦，后猛，肖满秋，李臣，潘皓，张允刚，卢凌志，侯隆英，葛瑞华，吴问胜，李强. 2023. AP2/ERF转录因子参与植物次生代谢和逆境胁迫响应的研究进展. 植物遗传资源学报，24 (5)：1223–1235.
[12]	Li W，Geng Z W，Zhang C P，Wang K L，Jiang X Q. 2021. Whole-genome characterization of Rosa chinensis AP2/ERF transcription factors and analysis of negative regulator RcDREB2B in Arabidopsis. BMC Genomics，22：1–20.
[13]	Li Y，Zhang H，Zhang Q，Liu Q C，Zhai H，Zhao N，He S Z. 2019. An AP2/ERF gene，IbRAP2-12, from sweetpotato is involved in salt and drought tolerance in transgenic Arabidopsis. Plant Science，281：19–30.
[14]	Liu Zhiwei，Xiong Yangyang，Li Tong，Yan Yajun，Han HongRun，Wu Zhijun，Zhuang Jing. 2014. Isolation and expression profiles analysis of two ERF subfamily transcription factor genes under temperature stresses in Camellia sinensis. Plant Physiology Journal，50 (12)：1821–1832. (in Chinese)
	刘志薇，熊洋洋，李彤，严雅君，韩洪润，吴致君，庄静. 2014. 茶树中两个ERF类转录因子的分离及不同茶树中温度胁迫的响应分析. 植物生理学报，50 (12)：1821–1832.
[15]	Ma Z M，Hu L J，Jiang W Z. 2024. Understanding AP2/ERF transcription factor responses and tolerance to various abiotic stresses in plants：a comprehensive review. International Journal of Molecular Sciences，25 (2)：893–915.
[16]	Nakano T，Suzuki K，Fujimura T，Shinshi H. 2006. Genome-wide analysis of the ERF gene family in Arabidopsis and rice. Plant Physiology，140 (2)：411–432.
[17]	Okamuro J K.，Caster B，Villarroel R，Van M M，Jofuk K D. 1997. The AP2 domain of APETALA2 defines a large new family of DNA binding proteins in Arabidopsis. Proceedings of the National Academy of Sciences，94 (13)：7076–7081.
[18]	Sakuma Y，Liu Q，Dubouzet J G，Abe H，Shinozaki K，YamaguchiShinozaki K. 2002. DNA-binding specificity of the ERF/AP2 domain of Arabidopsis DREBs，transcription factors involved in dehydration-and cold-inducible gene expression. Biochemical and Biophysical Research Communications，290 (3)：998–1009.
[19]	Su L，Zhang Y C，Yu S，Geng L F，Lin S，Ouyang L，Jiang X Q. 2022. RcbHLH59-RcPRs module enhances salinity stress tolerance by balancing Na⁺/K⁺ through callose deposition in rose（Rosa chinensis）. Horticulture Research，10 (3)：uhac291.
[20]	Tang Zheng，Chen Sique，Xu Qian，Zhong Weijie，Liu Qing，Zhu Shiyang. 2024. Functional study of AP2/ERF in response to black rot at broccoli seedling stage. Acta Horticulturae Sinica，51 (11)：2523–2539. (in Chinese)
	唐征，陈思雀，徐谦，钟伟杰，刘庆，朱世杨. 2024. AP2/ERF在青花菜苗期响应黑腐病的功能研究. 园艺学报，51 (11)：2523–2539.
[21]	Wang Waner，Wang Bingzhen，Zhou Yan，Fu Jiabin，Chen Lingling，Niu Yiding. 2023. Analysis of expression characteristics and cold resistance function of MfERF028 gene in the AP2/ERF family of Medicago falcata. Chinese Journal of Grassland，45 (6)：15–22. (in Chinese)
	王婉儿，王炳臻，周艳，付佳宾，陈玲玲，牛一丁. 2023. 黄花苜蓿AP2/ERF家族MfERF028基因的表达特性与耐寒功能分析. 中国草地学报，45 (6)：15–22.
[22]	Wei X R，Ling W，Ma Y W，Du J L，Cao F X，Chen H X，Chen J R，Li Y F. 2022. Genome-wide analysis of the trehalose-6-phosphate synthase gene family in rose（Rosa chinensis）and differential expression under heat stress. Horticulturae，8 (5)：429–446.
[23]	Xu H，Wang X C，Chen J. 2010. Overexpression of the Rap2.4f transcriptional factor in Arabidopsis promotes leaf senescence. Science China Life Sciences，53 (10)：1221–1226.
[24]	Yao Y，He R J，Xie Q L，Zhao X H，Deng X M，He J B，Song L L，He J，Marchant A，Chen X Y. 2017. Ethylene response factor 74（ERF74）plays an essential role in controlling a respiratory burst oxidase homolog D（RbohD）‐dependent mechanism in response to different stresses in Arabidopsis. New Phytologist，213 (4)：1667–1681.
[25]	Yue Manfang，Zhang Chun，Wu Zhongyi. 2022. Research progress in the structural and functional analysis of plant transcription factor AP2/ERF protein family. Biotechnology Bulletin，38 (12)：11–26. (in Chinese)
	悦曼芳，张春，吴忠义. 2022. 植物转录因子AP2/ERF家族蛋白结构和功能的研究进展. 生物技术通报，38 (12)：11–26.
[26]	Zhao Yabin，Zhang Bo，Hu Lin，Yuan Hao，Zhu Junyong，Sun Lifang，Zhang Wenhui. 2018. Cloning and expression analysis of BpERF1 and BpERF2 genes from Betula platyphylla. Journal of Sichuan Agricultural University，36 (3)：329–336. (in Chinese)
	赵雅彬，张博，胡林，袁浩，朱俊勇，孙丽芳，张文慧. 2018. 白桦BpERF1和BpERF2基因的克隆与表达模式分析. 四川农业大学学报，36 (3)：329–336.