《植物生理学报》 2016, 52(7): 1099-1109

油茶2个甘油-3-磷酸酰基转移酶基因的克隆与组织表达特异性分析

魏琦琦, 赵广, 何潇, 龙洪旭, 李泽, 谭晓风, 张琳^*

中南林业科技大学, 经济林培育与保护省部共建教育部重点实验室, 经济林育种与栽培国家林业局重点实验室, 经济林培育与利用湖南省2011协同创新中心, 长沙410004

通信作者：张；E-mail: triwoodtim918@126.com

摘　要：

甘油-3-磷酸酰基转移酶(glycerol-3-phosphate acyltransferase, GPAT)是三酰甘油生物合成Kennedy途径中的关键酶, 在植物的油脂合成与抗逆等方面起着重要作用。本研究在转录组数据基础上, 通过RACE技术, 从我国重要木本食用油料树种油茶中克隆了2个GPAT基因CoATS1和CoGPAT6的全长cDNA序列, 编码序列长度分别为1 353和1 488 bp, 分别编码450和495个氨基酸。蛋白结构预测发现2个蛋白C-端都含有1个酰基转移酶结构域, CoGPAT6的N-端还具有1个类卤酸脱卤酶结构域。多序列比对和系统进化分析结果表明, CoATS1与茶树的CsGPAT聚为一类, CoGPAT6与芝麻的SiGPAT6聚为一类。实时荧光定量PCR分析表明, CoATS1和CoGPAT6基因在检测的各样本中均有表达, 其中CoATS1在叶片中的表达量显著高于其他组织, 在种子的发育过程中, 表达呈递减变化; CoGPAT6在花瓣中表达量最高, 在种子中呈“降低-升高-降低”的表达特征。本研究为阐明GPAT类基因在油茶种子油脂合成途径中的功能奠定了基础。

关键词：油茶; 甘油-3-磷酸酰基转移酶; 蛋白结构; 系统进化树; 表达分析

收稿：2016-05-06   修定：2016-06-16

资助：国家自然科学基金(31470684)。

Cloning and expression analysis of two glycerol-3-phosphate acyltransferase (GPAT) genes in oil-tea tree (Camellia oleifera)

WEI Qi-Qi, ZHAO Guang, HE Xiao, LONG Hong-Xu, LI Ze, TAN Xiao-Feng, ZHANG Lin^*

Key Laboratory of Cultivation and Protection for Non-Wood Forest Trees, Ministry of Education, Key Lab of Non-Wood Forest Products of State Forestry Administration, Cooperative Innovation Center of Cultivation and Utilization for Non-Wood Forest Trees of Hunan Province, Central South University of Forestry and Technology, Changsha 410004, China

Corresponding author: ZHANG Lin; E-mail: triwoodtim918@126.com

Abstract:

As a functionally important enzyme in the Kennedy pathway involved in triacylgycerol (TAG) accumulation, glycerol-3-phosphate acyltransferase (GPAT) plays a key role in lipid biosynthesis and tolerance to abiotic stress in plants. Based on Camellia oleifera transcriptome data obtained before, we cloned two fulllength cDNA of GPAT, i.e. CoATS1 and CoGPAT6 via RACE method. The coding sequences of the two cDNA are 1 353 and 1 488 bp in length, encoding 450 and 495 amino acids, respectively. Prediction of protein structure showed that both CoATS1 and CoGPAT6 display an acyltransferase domain (AT domain), and CoGPAT6 has a haloacid dehalogenase-like hydrolase (HAD-like) domain as well. Protein multiple sequence alignment and phylogenetic analysis showed that chloroplast GPATs and endoplasmic reticulum-GAPT6s are divided into two different groups. On the phylogenetic tree, CoATS1 clustered with CsGAPT of Camellia sinensis, and CoGPAT6 clustered with SiGPAT6 of Sesamum indicum. qRT-PCR analysis indicated that both CoATS1 and CoGPAT6 expressed in all of the tissues used in this study. The expression level of CoATS1 in leaf was significantly higher than that in the other tissues and it gradually decreased in the developing seeds. CoGPAT6 had a highest expression amount in petal and exhibited a pattern of “decrease first, then increase and decrease finally”. This study provides an important basis for uncovering the detailed role of CoATS1 and CoGPAT6 in lipid biosynthesis in seeds of C. oleifera.

Key words: Camellia oleifera; glycerol-3-phosphate acyltransferase; protein structure; phylogenetic tree; expression analysis