《植物生理学报》 2018, 54(6): 1073-1084

本氏烟PP2A亚基基因的表达模式和克隆

陈孝仁^*, 黄沈鑫, 张烨, 李艳朋, 车通, 纪兆林

扬州大学园艺与植物保护学院, 江苏扬州225009

通信作者：陈孝仁；E-mail: xrchen@yzu.edu.cn

摘　要：

蛋白磷酸酶2A (protein phosphatase 2A, PP2A)由结构亚基A、催化亚基C和调节亚基B组成, 控制生物体内多种生命代谢过程, 但目前尚不清楚其在植物抗病中的作用。本研究利用拟南芥、普通烟草、番茄和水稻PP2A的亚基序列, 比对分析获得本氏烟(Nicotiana benthamiana)中对应的亚基序列。利用实时定量RT-PCR技术, 分析了6个亚基基因在辣椒疫霉侵染本氏烟阶段(灌根后8、24、72 h)以及2个结构亚基基因在本氏烟不同组织部位(根、茎、叶、花和种子)中的转录水平。结果发现, 在病原物侵染时, 多数基因均显著上调表达, NbPP2Ac-4在侵染阶段的表达量显著高于其他基因; 2个结构亚基基因(NbPP2Aa-1和NbPP2Aa-2)在茎、叶、花和种子中的表达量均显著高于在根部中的表达量。克隆获得本氏烟和辣椒中2个结构亚基基因的全长cDNA序列, 它们的二级结构主要为α和η螺旋, 含有保守的功能域, 与其他植物和人的结构亚基具有很高的同源性。

关键词：蛋白磷酸酶2A; 植物抗病性; 转录水平; 功能域; 辣椒疫霉

收稿：2018-01-04   修定：2018-05-31

资助：国家自然科学基金(31671971)、扬州市自然科学基金(YZ2016121)、公益性行业(农业)科研专项(201303018)和2016年度扬州大学优秀青年骨干教师培养对象。

Expression profile and cloning of genes encoding PP2A subunits of Nicotiana benthamiana

CHEN Xiao-Ren^*, HUANG Shen-Xin, ZHANG Ye, LI Yan-Peng, CHE Tong, JI Zhao-Lin

College of Horticulture and Plant Protection, Yangzhou University, Yangzhou, Jiangsu 225009, China

Corresponding author: CHEN Xiao-Ren; E-mail: xrchen@yzu.edu.cn

Abstract:

Ser/Thr protein phosphatase 2A (PP2A), consisting of a catalytic subunit C, a scaffold subunit A, and a highly variable regulatory subunit B, plays important roles in cellular processes in organisms. However, its role in plant disease resistance is obscure. In the present study, the genes encoding PP2A subunits of Nicotiana benthamiana were obtained by searching its genome database using Arabidopsis thaliana, common tobacco, tomato and rice PP2A subunit sequences as queries. Real-time RT-PCR was performed to determine the expression pattern of six selected genes during an infection period of N. benthamiana by Phytophthora capsici (0, 8, 24, 72 h post-inoculation), and of two PP2A-A genes in different plant tissues (root, stem, leaf, flower and seed). It turned out that most of the genes demonstrated significant up-regulation during the infection stage relative to the un-infected control. The most significant up-regulation was observed for NbPP2Ac-4 during the infection course. PP2A-A genes (NbPP2Aa-1 and NbPP2Aa-2) showed significantly higher expression level in stem, leaf, flower and seed than in root. These two PP2A-A genes were cloned with full length from N. benthamiana and its relative bell pepper. Sequence analysis showed that α- and η-helices were the main components of their secondary structure. They possess conserved domains such as HEAT_2 and showed high similarity to the homologues of PP2A-A in other plant species and human. This study provides essential experimental data for elucidating the roles of PP2A-A subunit in plant resistance against diseases.

Key words: protein phosphatase 2A; plant disease resistance; transcriptional level; domain; Phytophthora capsici