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Journal of Integrative Agriculture  2024, Vol. 23 Issue (12): 4032-4045    DOI: 10.1016/j.jia.2023.11.013
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Mepiquat chloride increases the Cry1Ac protein content of Bt cotton under high temperature and drought stress by regulating carbon and amino acid metabolism

Dian Jin*, Yuting Liu*, Zhenyu Liu, Yuyang Dai, Jianing Du, Run He, Tianfan Wu, Yuan Chen, Dehua Chen#, Xiang Zhang#

Jiangsu Key Laboratory of Crop Genetics and Physiology/Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University, Yangzhou 225009, China
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摘要  

本文研究了高温干旱胁迫下甲哌鎓(DPC)对BtCry1Ac蛋白含量的影响,以期为Bt棉育种和高产高效栽培提供理论参考。于2020年和2021年在扬州大学农牧场进行,以“泗抗3号”为材料,采用盆栽栽培。供试材料在人工气候室高温和干旱胁迫后,进行喷施20 mg L-1 DPC和清水(CK)处理。处理7d后,测定Cry1Ac蛋白含量、α-酮戊二酸含量、丙酮酸含量、谷氨酸合成酶活性、谷氨酸草酰乙酸转氨酶活性、可溶性蛋白含量和氨基酸含量,并进行转录组测序。结果表明,与CK相比,DPC处理Cry1Ac蛋白含量增加了4.7%-11.9%α-酮戊二酸含量、丙酮酸含量、谷氨酸合成酶活性、谷氨酸草酰乙酸转氨酶活性、可溶性蛋白含量和氨基酸含量均有所增加。转录组分析显示, DPC处理7542个基因上调,10449个基因下调。进一步分析表明,差异表达基因主要涉及氨基酸代谢和碳代谢等过程。其中编码6-磷酸果糖激酶、丙酮酸激酶、谷氨酸丙酮酸转氨酶、丙酮酸脱氢酶、柠檬酸合成酶、异柠檬酸脱氢酶、2-氧代戊二酸脱氢酶的基因显著上调,谷氨酸合成酶、1-吡咯啉-5-羧酸脱氢酶、谷氨酸草酰乙酸转氨酶、N-乙酰谷氨酸合成酶和乙酰鸟氨酸脱乙酰酶也显著上调。说明DPC提高糖酵解途径、柠檬酸循环的运行速率,增加了丙酮酸、α-酮戊二酸和草酰乙酸。DPC处理还显著上了调谷氨酸合成酶和吡咯烷-5-羧酸脱氢酶的编码基因,下调谷氨酰胺合成酶的编码基因,上调谷氨酸草酰乙酸转氨酶和N-乙酰谷氨酸合成酶的编码基因。综上,DPC处理通过调节高温干旱胁迫后Bt棉自身碳和氨基酸代谢能力,增强天门冬氨酸、谷氨酸、丙酮酸和精氨酸含量,进而为Cry1Ac蛋白合成提供充足原料,并提高其含量。



Abstract  

The effects of mepiquat chloride (DPC) on the Cry1Ac protein content in Bacillus thuringiensis (Bt) cotton boll shells under high temperature and drought stress were investigated to provide a theoretical reference for Bt cotton breeding and high-yield and -efficiency cotton cultivation.  This study was conducted using Bt cotton cultivar ‘Sikang 3’ during the 2020 and 2021 growing seasons at Yangzhou University Farm, Yangzhou, Jiangsu Province, China.  Potted cotton plants were exposed to high temperature and drought stress, and sprayed with either 20 mg L−1 DPC or water (CK).  Seven days after treatment, the Cry1Ac protein content, α-ketoglutarate content, pyruvic acid content, glutamate synthase activity, glutamic oxaloacetic transaminase activity, soluble protein content, and amino acid content were measured, and transcriptome sequencing was performed.  DESeq was used for differential gene analysis.  Under the DPC treatment, the Cry1Ac protein content increased by 4.7–11.9% compared to CK.  The α-ketoglutarate content, pyruvic acid content, glutamate synthase activity, glutamic oxaloacetic transaminase activity, soluble protein content, and amino acid content all increased.  Transcriptome analysis revealed 7,542 upregulated genes and 10,449 downregulated genes for DPC vs. CK.  Gene ontology (GO) and Kyoto Encyclopedia of Gene and Genomes (KEGG) analyses showed that the differentially expressed genes were mainly involved in biological processes, such as carbon and amino acid metabolism.  For example, genes encoding 6-phosphofructokinase, pyruvate kinase, glutamic pyruvate transaminase, pyruvate dehydrogenase, citrate synthase, isocitrate dehydrogenase, 2-oxoglutarate dehydrogenase, glutamate synthase, 1-pyrroline-5-carboxylate dehydrogenase, glutamic oxaloacetic transaminase, amino-acid N-acetyltransferase, and acetylornithine deacetylase were all significantly upregulated.  The DPC treatment increased pyruvate, α-ketoglutarate, and oxaloacetate by increasing the operational rate of the glycolytic pathway of the citric acid cycle.  It also significantly upregulated the genes encoding glutamate synthase, pyrrolidine-5-carboxylic acid dehydrogenase, glutamate oxaloacetate transaminase, and N-acetylglutamate synthetase, while it downregulated the genes encoding glutamine synthetase.  Therefore, the synthesis of aspartic acid, glutamic acid, pyruvate, and arginine increased after treatment with DPC, and the Cry1Ac protein content was increased by regulating carbon and amino acid metabolism.

Keywords:  amino acid metabolism       Bt cotton        carbon metabolism        Cry1Ac protein        mepiquat chloride  
Received: 06 May 2023   Accepted: 12 October 2023
Fund: 
This work was supported by the National Natural Science Foundation of China (31901462), the Natural Science Foundation of the Jiangsu Higher Education Institutions, China (22KJA210005), the Priority Academic Program Development of Jiangsu Higher Education Institutions, China (PAPD), and the Brand Professional Construction Program of Jiangsu Higher Education Institutions, China.  
About author:  Dian Jin, Tel: +86-514-87979357, E-mail: 2774134783@qq.com; Yuting Liu, Tel: +86-514-87979357, E-mail: 2544223887@qq.com; #Correspondence Xiang Zhang, Tel: +86-514-87979357, E-mail: yzzhangxiang@163.com; Dehua Chen, Tel: +86-514-87979357, E-mail: cdh@yzu.edu.cn * These authors contributed equally to this study.

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