《植物生理学报》 2017, 53(8): 1399-1408

小桐子中一个新的ERF家族转录因子JcERF3基因的克隆及功能分析

王莎莎^*, 赵洪春^*, 邓凤飞, 龚明^**

云南师范大学生命科学学院, 生物能源持续开发利用教育部工程研究中心, 云南省生物质能与环境生物技术重点实验室, 昆明650500

通信作者：王莎莎；E-mail: gongming63@163.com

摘　要：

从小桐子(Jatropha curcas)幼苗的叶片组织中分离出一个新的AP2/ERF类转录因子的编码基因, 命名为JcERF3。序列分析显示其完整编码区为729 bp, 编码蛋白由242个氨基酸组成。推测的蛋白质与其他物种的ERF3蛋白具有很高的序列相似性, 并含有一个保守的AP2结构域。对启动子元件的预测显示存在一个乙烯响应元件(ERE), 暗示JcERF3的表达可能受到乙烯的调节。核定位信号预测JcERF3可能定位于细胞核。过表达JcERF3的转基因烟草(Nicotiana tabacum)离体叶盘在200 mmol•L^-1 NaCl溶液中的失绿程度显著低于野生型。经0°C低温胁迫24 h后, 转基因株系中游离脯氨酸和可溶性糖含量为野生型的3.79和1.67倍以上, 而电解质渗漏率仅为野生型的74%左右。12°C低温下转基因株系T1代种子的萌发速度和萌发率也远高于野生型。综上可知, JcERF3作为一个新的AP2/ERF转录因子, 其过表达赋予转基因烟草对低温和盐胁迫较强的耐受性, 为小桐子的抗逆遗传改良提供了重要的基因资源。

关键词：小桐子; 转录因子ERF; 过量表达; 低温胁迫; 盐胁迫

收稿：2017-06-09   修定：2017-06-18

资助：国家自然科学基金(31460182和31460059)。

Isolation and functional characterization of a new ERF transcription factor JcERF3 from Jatropha curcas

WANG Sha-Sha^*, ZHAO Hong-Chun^*, DENG Feng-Fei, GONG Ming^**

School of Life Sciences, Yunnan Normal University; Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of Education; Key Laboratory of Biomass Energy and Environmental Biotechnology of Yunnan Province, Kunming 650500, China

Corresponding author: WANG Sha-Sha; E-mail: gongming63@163.com

Abstract:

A novel candidate for ERF-like transcription factor was isolated from the leaf tissue of Jatropha curcas, and designated as JcERF3. The complete coding region (CDS) of JcERF3 was 729 bp, encoding a polypeptide of 242 amino acid residues. The predicted JcERF3 protein shares high identities with ERF3-like proteins from other plant species, and containes a conserved DNA-binding domain (AP2/ERF domain) with 58 amino acids. An ethylene-responsive element (ERE) was predicted as a cis-element in the promoter region, indicating the potential regulation of ethylene on expression of JcERF3. Prediction of nuclear localization sequence (NLS) implies that JcERF3 protein probably localizes in nucleus. Overexpression of JcERF3 conferred salinity tolerance in transgenic tobacco, as was evident by leaf disk assays. When inoculated in the solution of 200 mmol•L^-1 NaCl, leaf disks of transgenic plants maintained a relatively higher content of chlorophyll, as compared to the wild type (WT). Beyond that, when exposed to chilling stress (0°C) for 24 h, the transgenic plants showed increased proline (at least 3.79-fold) and soluble sugar (at least 1.67-fold) contents and reduced electrolyte leakage (approx. 74%), as compared to WT. Germination assays of T1 seeds under chilling stress also indicates the transgenic tobacco seeds showed earlier and higher percent of germination, as compared to WT. Taken together, as a novel AP2/ERF transcription factor, overexpression of JcERF3 conferred transgenic tobacco plants an enhanced tolerance to chilling and high salinity stresses. It can be used as a potential candidate gene for genetic improvement of stress tolerance in J. curcas.

Key words: Jatropha curcas; transcription factor ERF; overexpression; chilling stress; salinity stress