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薏苡VQ4基因的克隆及耐盐性初步分析

王玉娇1，王永乐1，添长久1，郁春旺1，吕佳斌2，朱加保1,*   

  1. 1 安徽省农业科学院经济作物研究所, 安徽合肥 230001; 2安徽农业大学林学与园林学院, 安徽合肥 230036
  • 收稿日期:2025-04-27 修回日期:2025-09-10 接受日期:2025-09-10 网络出版日期:2025-09-24
  • 通讯作者: 朱加保, E-mail: 13955611798@139.com
  • 基金资助:
    本研究由国家自然科学基金青年项目(3240151950)资助。

Cloning of the VQ4 gene and preliminary analysis of its role in salt tolerance in Coix lacryma-jobi L.

WANG Yu-Jiao1,WANG Yong-Le1,TIAN Chang-Jiu1,YU Chun-Wang1,LYU Jia-Bin2,ZHU Jia-Bao1,*   

  1. 1 Institute of Industrial Crops, Anhui Academy of Agricultural Sciences, Hefei 230001, Anhui, China; 2 School of Forestry and Landscape Architecture, Anhui Agricultural University, Hefei 230036, Anhui, China
  • Received:2025-04-27 Revised:2025-09-10 Accepted:2025-09-10 Published online:2025-09-24
  • Contact: 朱加保, E-mail: 13955611798@139.com
  • Supported by:
    This study was supported by the Young Scientists Fund of the National Natural Science Foundation of China (3240151950).

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摘要：

VQ蛋白是一类含有“VQ”保守结构域的植物特异性蛋白家族，在非生物胁迫响应和生长发育中的重要作用备受关注。本研究以“皖薏2号”为材料，利用生物信息学分析手段、荧光定量PCR技术、亚细胞定位、酵母双杂试验等对薏苡VQ4进行分子特征鉴定；通过酵母异源表达和过表达拟南芥验证其对盐胁迫的响应。结果表明，ClVQ4的开放阅读框长度为594 bp，编码197个氨基酸，并且ClVQ4是等电点为6.43的不稳定亲水性蛋白。顺式作用元件分析结果发现，ClVQ4基因启动子序列含有多个激素响应元件和逆境胁迫响应元件。ClVQ4基因的表达受到茉莉酸甲酯(MeJA)和脱落酸(ABA)的诱导，其表达量在盐胁迫下显著上调。亚细胞定位和酵母双杂结果表明，ClVQ4蛋白主要定位于细胞核和细胞膜中，并且可以和薏苡WRKY30相互作用，同时可以和其他VQ蛋白相互作用形成异源二聚体。在NaCl处理下，酿酒酵母异源表达ClVQ4基因可以提高酵母的存活率。在盐胁迫下，ClVQ4过表达拟南芥的萌发率和根长均显著高于野生型，且各转基因株系中POD、SOD和CAT活性均显著高于野生型，而MDA含量显著低于野生型。

关键词: 薏苡, ClVQ4, 基因克隆, 亚细胞定位, 耐盐性

Abstract:

VQ proteins, a plant-specific protein family characterized by a conserved VQ domain, have garnered considerable attention due to their critical roles in abiotic stress responses as well as plant growth and development. In this study, the Coix lacryma-jobi L. cultivar ‘Wanyi 2’ was used as the experimental material. A combination of bioinformatics analysis, quantitative real-time PCR, subcellular localization, and yeast two-hybrid assays was employed to characterize the molecular properties of ClVQ4. Its function in salt stress response was further investigated through heterologous expression in yeast and genetic transformation in Arabidopsis thaliana. The results revealed that ClVQ4 contains an open reading frame (ORF) of 594 bp, encoding a 197-amino-acid protein predicted to be an unstable, hydrophilic protein with an isoelectric point (pI) of 6.43. Cis-acting element analysis of the ClVQ4 promoter identified multiple hormone- and stress-responsive elements. The expression of ClVQ4 was induced by methyl jasmonate (MeJA) and abscisic acid (ABA), and significantly upregulated under salt stress. Subcellular localization and yeast two-hybrid assays showed that ClVQ4 is mainly localized in the nucleus and cell membrane, interacts with Coix WRKY30, and can form heterodimers with other VQ proteins. Under NaCl treatment, the yeast strain expressing ClVQ4 exhibited enhanced survival. Furthermore, overexpression of ClVQ4 in Arabidopsis resulted in higher germination and survival rates and longer root lengths compared to the wild type (WT) under salt stress. Additionally, transgenic lines exhibited significantly higher activities of antioxidant enzymes (POD, SOD, and CAT) and significantly lower malondialdehyde (MDA) content than WT, suggesting a positive regulatory role of ClVQ4 in salt stress tolerance.

Key words: Coix lacryma-jobi L., ClVQ4, gene cloning, subcellular localization, salt tolerance

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