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| Screening and Validation of Upstream Regulatory Proteins of Cinnamomum bodinieri CbADH Gene |
| ZHANG Nan, HAN Hao-Zhang*, ZHANG Li-Hua, ZHAO Rong, LI Su-Hua, WANG Fang |
| Institute of Biology and Materials, Suqian University, Suqian 223800, China |
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Abstract Respiratory metabolism is closely related to plant tolerance to saline and alkali stress, and ethanol dehydrogenase is a key enzyme for anaerobic respiration in plants, which is up-regulated under saline and alkali stress and plays an important role in plant tolerance to saline and alkali stress. In order to clarify the molecular mechanism of CbADH gene involved in the salinity stress tolerance of Cinnamomum bodinieri, in this study, the promoter sequence of CbADH gene was cloned using the alkali-tolerant Cinnamomum bodinieri line as the material, and the upstream regulatory factors of CbADH gene was screened out by yeast one-hybridization, and the function of the upstream regulatory proteins on the promoter of CbADH gene was verified by the dual luciferase reporter gene technique. The results showed that the length of the obtained CbADH gene promoter fragment was 1 996 bp; the CbADH gene promoter fragment contained light-responsive regulatory elements, ABA-responsive elements, phloem-expression-responsive elements, MeJA-responsive elements, GA-responsive elements, stress-responsive elements, SA response element, healing tissue response element, transcription factor MYB binding site, transcription factor bHLH binding site, and transcription factor WRKY binding site. A total of 17 successfully annotated reciprocal proteins were obtained by yeast one-hybrid technique, mainly including protein kinase superfamily, amino acid transporters, mediator of ABA-regulated dormancy 1 (MARDⅠ) zinc finger structural proteins, Arabidopsis thaliana homeobox 6 (ATHB-6) proteins, trisaccharide phosphoribosylceramide isomerase proteins, late embryogenesis abundant protein (LEA) proteins, tyrosine/dopamine decarboxylases, mannitol dehydrogenase proteins, ATP synthases, and cytochrome B5 family proteins. According to the results of the dual luciferase reporter gene experiment technology, it was shown that the CbATHB6 protein activated the promoter activity of CbADH gene in plants. Based on these findings, the transcription factor CbATHB6 could bind to the promoter sequence of the CbADH gene and respond to salt-alk stress by regulating the expression of the CbADH gene. This study provides a basis for analyzing the molecular mechanism of respiratory metabolism of Cinnamomum bodinieri in response to saline and alkaline stress.
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Received: 25 March 2025
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Corresponding Authors:
*21011@squ.edu.cn
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[1] 鲍庆晗. 2023. 藜麦响应盐碱胁迫的生理特性及转录组与代谢组机制解析[D]. 博士学位论文, 内蒙古农业大学, 导师: 张永平, pp.91-97.
(Bao Q H.2023. Physiological characteristics and mechanism analysis of transcriptome and metabolome of quinoa under salt and alkali stress[D]. Thesis for Ph. D., Inner Mongolia Agricultural University, Supervisor: Zhang Y P, pp. 91-97.)
[2] 陈运梁, 邹竹荣, 杨双龙. 2023. 外源茉莉酸甲酯对盐胁迫下小桐子幼苗渗透调节和脯氨酸代谢的影响[J]. 西北植物学报, 43(5): 794-804.
(Chen Y L, Zou Z R, Yang S L.2023. Effect of exogenous methyl jasmonate on ssmotic adjustment capacity and proline metabolism of Jatropha curcas seedlings under salt stress[J]. Acta Botanica Boreali-Occidentalis Sinica, 43(5): 794-804.)
[3] 陈素梅, 王新慧, 李菲, 等. 2022. ABA调控植物盐胁迫应答机制研究进展[J]. 南京农业大学学报, 45(5): 856-863.
(Chen S M, Wang X H, Li F, et al.2022. Advances in ABA regulated responses of plant to salt stress[J]. Journal of Nanjing Agricultural University, 45(5): 856-863.)
[4] 程斌, 张创娟, 杨乐, 等. 2022. 绿豆乙醇脱氢酶基因生物信息学分析及其在镉胁迫下的表达特性变化[J]. 植物资源与环境学报, 31(02): 10-21.
(Cheng B, Zhang C J, Yang L,et al.2022. Bioinformatics analysis on alcohol dehydrogenase genes in Vigna radiata and changes of their expression characteristics under cadmium stress[J]. Journal of Plant Resources and Environment, 31(02): 10-21.)
[5] 郭家鑫. 2023.不同盐碱胁迫对棉花苗期蛋白质组和代谢组的影响[D]. 硕士学位论文, 石河子大学, 导师: 闵伟, pp. 78-81.
(Guo J X.2023. Effects of different salt and alkali stress on proteome and metabolome of cotton seedlings[D]. Thesis for M.S., Shihezi University, Supervisor: Yan W, pp. 78-81.)
[6] 焦龙, 陈勋, 谭荣荣, 等. 2023. 植物茉莉酸与水杨酸信号途径互作研究进展[J]. 植物生理学报, 59(8): 1489-1504.
(Jiao L, Chen X, Tan R R, et al.2023. Research progress on the interaction between jasmonic acid and salicylic acid signaling pathways in plants[J]. Plant Physiology Journal, 59(8): 1489-1504.)
[7] 李俊伟, 刘景辉, 王俊英, 等. 2022. 盐与碱胁迫对燕麦离子平衡和有机酸含量的影响[J]. 西北植物学报, 42(10): 1700-1710.
(Li J W, Liu J H, Wang J Y, et al.2022. Effect of salt and alkali stress on oat ion balance and organic acid content[J]. Acta Botanica Boreali-Occidentalia Sinica, 42(10): 1700-1710.)
[8] 沙向红. 2015. 盐胁迫对玉米幼苗根系乙醇脱氢酶与乙醛脱氢酶表达的影响[J]. 贵州农业科学, 43(03): 47-50.
(Sha X H.2015. Effect of salt stress on expression of ethanol dehydrogenase (ADH2) and acetaldehyde dehydrogenase (ALDH2) in roots of young corn seedlings[J]. Guizhou Agricultural Sciences, 43(03): 47-50.)
[9] 苏炜华, 黄宁, 凌辉, 等. 2017. 甘蔗乙醇脱氢酶基因的克隆与表达[J]. 应用与环境生物学报, 23(03): 474-481.
(Su W H, Hang N, Ling H, et al.2017. Cloning and expression of ScADH from sugarcane[J]. Chinese Journal of Applied and Environmental Biology, 23(03): 474-481.)
[10] 魏云竹. 2017. 拟南芥乙醇脱氢酶ADH的抗病性功能研究[D]. 硕士学位论文, 兰州大学, 导师: 安黎哲, pp. 22-25.
(Wei Y Z.2017. The function of disease resistance of alcohol dehydrogenase in Arabidopsis thaliana[D]. Thesis for M.S., Lanzhou University, Supervisor: An L Z, pp. 22-25.)
[11] 武亮亮, 姚磊, 马瑞, 等. 2016. 马铃薯HD-ZipⅠ家族ATHB12基因的克隆及功能鉴定[J]. 作物学报, 42(08): 1112-1121.
(Wu L L, Yao L, Ma R, et al.2016. Cloning and functional identification of the ATHB12 gene of HD-ZipⅠ family in potato (Solanum tuberosum L.)[J]. Acta Agronomica Sinica, 42(08): 1112-1121.)
[12] 徐忠山. 2021. 利用多组学联合分析燕麦响应干旱胁迫及盐胁迫的分子机制[D]. 博士学位论文, 内蒙古农业大学, 导师: 逯晓萍, pp. 96-98.
(Xu Z S.2021.Using multi-omics to analyze the molecular mechanism of oat in response to drought stress and salt stress[D]. Thesis for Ph. D., Inner Mongolia Agricultural University, Supervisor: Lu X P, pp. 96-98.)
[13] 张恩平, 张淑红, Cordewener J, 等. 2005. 盐胁迫下不同盐敏感型番茄在蛋白质表达上的差异[J]. 沈阳农业大学学报, 36(01): 25-28.
(Zhang E P, Zhang S H, Cordewener J, et al.2005. Protein expressing difference in salt-tolerant and salt-sensitive tomato under salt stress[J]. Journal of Shenyang Agricultural University, 36(01): 25-28.)
[14] 周岩, 何男男, 魏琦超, 等. 2010. 同源异型域-亮氨酸拉链蛋白研究进展[J]. 湖北农业科学, 49(11): 2913-2917.
(Zhou Y, He N N, Wei Q C, et al.2010. Research progress in HD-Zip protein[J]. Hubei Agricultural Sciences, 49(11): 2913-2917.)
[15] Abe H, Urao T, Ito T, et al.2003. Arabidopsis AtMYC2(bHLH) and AtMYB2(MYB) function as transcriptional activators in abscisic acid signaling[J]. The Plant Cell, 15: 63-78.
[16] Conley T R, Peng H P, Shih M C.1999. Mutations afecting induction of glycolytic and fermentative genes during germination and environmental stresses in Arabidopsis[J]. Plant Physiology, 119(2): 599-608.
[17] Fang S, Hou X, Liang X.2021. Response mechanisms of plants under saline-alkali stress[J]. Frontiers in Plant Science, 12: 667458.
[18] Han H Z, Zhang L H, Li S H, et al.2023. Transcriptome and metabolome integrated analysis reveals the mechanism of cinnamomum bodinieri root response to alkali stress[J]. Plant Molecular Biology Reporter, 41(3): 470-488.
[19] Jiao P, Jiang Z Z, Wei X T, et al.2022. Overexpression of the homeobox-leucine zipper protein ATHB-6 improves the drought tolerance of maize (Zea mays L.)[J]. Plant Science, 316: 111159.
[20] Kursteiner O, Dupuis I, Kuhlemeier C.2003. The pyruvate decarboxylase 1 gene of Arabidopis is required during anoxia but not other environmental sresses[J]. Plant Physiol, 132: 968-978.
[21] Zhang H, Wang S, Li O, et al.2024. Genome-wide identification of alcohol dehydrogenase (ADH) gene family in oilseed rape (Brassica napus L.) and BnADH36 functional verification under salt stress[J]. BMC Plant Biology, 24(1): 1013.
[22] Zhang J, Yang D S, Li M X, et al.2016. Metabolic profiles reveal changes in wild and cultivated soybean seedling leaves under salt stress[J]. PLOS ONE, 11(7): e0159622.
[23] Zhou J, Qi A G, Wang B Q, et al.2022. Integrated analyses of transcriptome and chlorophyll fluorescence characteristics reveal the mechanism underlying saline-alkali stress tolerance in Kosteletzkya pentacarpos[J]. Frontiers in Plant Science, 13: 865572. |
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