油莎豆FAT家族全基因组鉴定及CeFATA的功能分析

doi:10.13560/j.cnki.biotech.bull.1985.2025-1395

摘要/Abstract

摘要：

目的酯酰-ACP硫酯酶（fatty acyl-ACP thioesterase, FAT）在植物油脂合成过程中发挥重要作用。解析油莎豆CeFAT家族基因功能，为高油油莎豆新品种的培育及油脂代谢改良提供理论依据。方法基于油莎豆全基因组数据，对CeFAT基因家族进行系统鉴定与序列分析，并对CeFATA蛋白进行亚细胞定位，通过酵母与烟草遗传转化体系对CeFATA进行功能分析。结果在油莎豆基因组中共鉴定到3个CeFATs基因成员CeFATA、CeFATB1和CeFATB2，3个成员具有相似的基因结构和理化性质，且分别位于3条染色体上。系统进化分析表明，CeFATA与水稻（Oryza sativa）OsFATA的亲缘关系最近，CeFATB1和CeFATB2与大豆（Glycine max）GmFATB的亲缘关系较近。实时荧光定量PCR结果显示，CeFATs基因在块茎5个不同发育时期均高水平表达，其中CeFATA的表达量最高。亚细胞定位结果证实CeFATA定位于叶绿体。将CeFATA在酿酒酵母中过表达结果显示，过表达酵母总脂含量比野生型提高了3.1%。底物偏好性试验表明CeFATA对C18:1具有选择性。CeFATA在烟草中过表达结果显示，过表达烟草叶片和种子总脂肪酸含量比野生型分别提高了4.74%和8.4%，同时淀粉含量均显著下降，可溶性糖含量呈现出在叶片中下降而种子中提高的趋势，蛋白质含量变化不显著。结论 CeFATA提高了酿酒酵母与烟草的总脂含量，促进质体中油酸的转运，使碳源从碳水化合物合成途径转向油脂合成途径。

关键词: 油莎豆, 酯酰-ACP硫酯酶, 亚细胞定位, 脂肪酸合成, 功能分析

Abstract:

Objective Fatty acyl-ACP thioesterase (FAT) plays a key role in plant oil synthesis. This study aimed to characterize FAT family genes in yellow nutsedge (Cyperus esculentus L.) and to provide a theoretical basis and technical support for breeding high-oil cultivars and improving lipid metabolism. Method Based on the whole-genome data of yellow nutsedge, the CeFAT gene family was systematically identified and subjected to sequence and phylogenetic analyses. The subcellular localization of CeFATA was determined, and the function of CeFATA was evaluated using yeast and tobacco genetic transformation systems. Result Three CeFAT genes, CeFATA, CeFATB1, and CeFATB2, were identified. They showed similar gene structures and physicochemical properties, and were located on three different chromosomes. Phylogenetic analysis indicated that CeFATA was most closely related to OsFATA from rice, whereas CeFATB1 and CeFATB2 were closely related to GmFATB from soybean. RT-qPCR analysis showed that CeFAT genes were highly expressed at five developmental stages of the tuber, with CeFATA showing the highest expression level. Subcellular localization confirmed that CeFATA was located in chloroplasts. Overexpression of CeFATA increased total lipid content in Saccharomyces cerevisiae by 3.1% and exhibited substrate preference for C18:1. In transgenic tobacco, CeFATA overexpression increased total fatty acid contents in leaves and seeds by 4.74% and 8.4%, accompanied by a significant decrease in starch content, a decrease soluble sugar content in leaves, an increase soluble sugar content in seeds, and no significant change in protein content. Conclusion CeFATA increases total lipid and oleic acid contents in yeast and tobacco, promotes the export of oleic acid from plastids, and redirects carbon flux from carbohydrate biosynthesis toward oil biosynthesis.

Key words: Cyperus esculentus L., fatty acyl-ACP thioesterase (FAT), subcellular localization, fatty acid biosynthesis, functional analysis

李淑婷, 王影, 丁家豪, 胡荣芳, 李琛, 宋亚楠, 薛金爱. 油莎豆FAT家族全基因组鉴定及CeFATA的功能分析[J]. 生物技术通报, 2026, 42(8): 162-172.

LI Shu-ting, WANG Ying, DING Jia-hao, HU Rong-fang, LI Chen, SONG Ya-nan, XUE Jin-ai. Genome-wide Identification of the FAT Gene Family and Functional Analysis of CeFATA in Cyperus esculentus[J]. Biotechnology Bulletin, 2026, 42(8): 162-172.

图/表 9

图 1 油莎豆CeFATs基因染色体定位（A）及与3种植物（拟南芥、大豆和水稻）共线性分析（B）

Fig. 1 Chromosomal localization of CeFATs genes in C. esculentus (A) and synteny analysis with three plant species (Arabidopsis, soybean, and rice) (B)

图2 CeFATs与不同物种FAT蛋白的系统进化树分析油莎豆：Ce；海枣：Pd；椰子：Cn；香蕉：Ma；野蕉：Mb；拟南芥：At；水稻：Os；大豆：Gm；向日葵：Ha；烟草：Nt；芝麻：Si；凤梨：Ac

Fig. 2 Phylogenetic tree analysis of CeFATs and FAT proteins from different speciesCe: Cyperus esculentusis; Pd: Phoenix dactylifera;Cn: Cocos nucifera; Ma: Musa acuminata; Mb: Musa balbisiana; At: Arabidopsis thaliana; Os: Oryza sativa; Gm: Glycine max; Ha: Helianthus annuus; Nt: Nicotiana tabacum; Si: Sesamum indicum;Ac: Ananas comosus

图3 油莎豆FATs家族蛋白结构与氨基酸构成分析

Fig. 3 Analysis of FAT family protein structure and amino acid composition in C. esculentus

图4 油莎豆CeFATs基因的启动子顺式作用元件分析

Fig. 4 Cis-acting element analysis of CeFATs genes in C. esculentus

图5 油莎豆CeFATs 在块茎不同发育时期的转录组表达谱分析（A）和RT-qPCR定量分析（B）不同小写字母表示存在显著差异（P<0.05）。下同

Fig. 5 Transcriptome expression profiling (A) and RT-qPCR analysis (B) of CeFATs at different tuber developmental stages in C. esculentusDifferent lowercase letters indicate significant differences (P<0.05). The same below

图6 CeFATA蛋白的亚细胞定位

Fig. 6 Subcellular localization of CeFATA protein

图7 转基因酵母总脂肪酸含量以及在外源添加脂肪酸下生长曲线A：总脂肪酸含量；B：外源添加4种脂肪酸下空载酵母生长曲线；C：外源添加4种脂肪酸下转基因酵母生长曲线

Fig. 7 Total fatty acid content and growth curve of transgenic yeast under exogenous fatty acid additionA: Total fatty acid content; B: Growth curves of empty-vector yeast with four exogenous fatty acids; C: Growth curves of transgenic yeast with four exogenous fatty acids

图8 转基因酵母脂肪酸组分含量

Fig. 8 Fatty acid contents of transgenic yeast

图9 转基因烟草株系生理生化指标A：叶片总脂含量。B：叶片蛋白含量。C：叶片淀粉含量。D：叶片可溶糖含量。E：种子总脂含量。F：种子蛋白含量。G：种子淀粉含量。H：种子可溶糖含量。WT：野生烟草；EV：转空载质粒烟草；CeFATA：转基因烟草株系。DW：干重

Fig. 9 Physiological and biochemical indices of transgenic tobacco linesA: Total lipid content in leaves. B: Protein content in leaves. C: Starch content in leaves. D: Soluble sugar content in leaves. E: Total lipid content in seeds. F: Protein content in seeds. G: Starch content in seeds. H: Soluble sugar content in seeds. WT: Wild tobacco; EV: Empty vector tobacco; CeFATA: Transgenic tobacco lines. DW: Dry weight

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