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Acta Agronomica Sinica ›› 2019, Vol. 45 ›› Issue (11): 1638-1648.doi: 10.3724/SP.J.1006.2019.94003

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Molecular mechanism of stearoyl-ACP desaturase gene FAB2 expression in peanut

LIU Hao,LU Qing,LI Hai-Fen,LI Shao-Xiong,CHEN Xiao-Ping,LIANG Xuan-Qiang(),HONG Yan-Bin()   

  1. Crops Research Institute, Guangdong Academy of Agricultural Science / Guangdong Provincial Key Laboratory of Crop Genetic Improvement, Guangzhou 510640, Guangdong, China
  • Received:2019-01-02 Accepted:2019-05-12 Online:2019-11-12 Published:2019-05-30
  • Contact: Xuan-Qiang LIANG,Yan-Bin HONG E-mail:liangxuanqiang@gdaas.cn;hongyanbin@gdaas.cn
  • Supported by:
    This study was supported by the National Natural Science Foundation of China(31771841);This study was supported by the National Natural Science Foundation of China(31801401);the Natural Science Foundation of Guangdong Province(2017A030311007);the National Key Research and Development Program(2018YFD0201009-02);Excellent PhD Program of Guangdong Academy of Agricultural Science

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Abstract:

Stearoyl-ACP desaturase, encoded by FAB2, and located on the upstream of oleic acid biosynthesis pathway, modulates the conversion of stearic acid (C18:0) into oleic acid (C18:1). The expression of FAB2 was increased at the early stage of seed development in high-oleic variety Kainong 176, but over-accumulated oleic acid repressed the FAB2 expression at the period of seed maturation. An F2 hybrid population was constructed using Kainong 176 and Kainong 70, showing that the content of oleic acid up to 60% directly repressed the FAB2 expression on the whole. The activity of peroxidase (POD) and content of ROS were increased at the early stage of seed development with the oleic acid gradual accumulation, but decreased at the maturation stage. Subcellular localization analysis indicated that FAB2 and FAD2 were located on chloroplast and endoplasmic reticulum, respectively. Encoding sequence polymorphism analysis of FAB2 suggested that amino acid deficient at the N-terminal of FAB2 protein sequence probably induced the high content of stearic acid in peanut. Furthermore, the promotor sequence of FAB2 contained multiple AT-rich region, and possessed the light responsive, hormone regulation, and transcription factor binding site cis-regulating elements. Taken together, this study found that over-accumulated oleic acid activated the POD-induced ROS robust pathway, and then regulated the expression of FAB2 by unknown transcription factor in nucleus. The results extend the knowledge of FAB2 function, and provide a relevant theoretical guidance in future peanut breeding for high oleic acid.

Key words: peanut, high oleic acid, FAB2, ROS, feedback regulation

Fig. 1

Oleic acid accumulation feedback regulates the expression level of FAB2 in peanut A: schematic diagram of oleic acid synthesis; B: photograph of Kainong 176 and Kainong 70; C: photograph of the six stages in the developing seeds; D: expression level of FAB2 during seeds development in the high- and normal-oleic varieties; E: contents of oleic acid and linoleic acid in Kainong 176 and Kainong 70; F: relative content changes of oleic acid and linoleic acid during seed development in Kainong 176 and Kainong 70."

Fig. 2

Oleic acid accumulated threshold value for repression of FAB2 expression in Peanut A: contents of oleic acid and linoleic acid in the individual plant of F2 population. OA: oleic acid; LOA: linoleic acid. B: relative expression of FAB2 in the individual plant of F2 population. F2-25 as the control, bar value represented three independent replications."

Fig. 3

Changed of POD activity and H2O2 content A: changes of POD activity; B: changes of H2O2 content; ** P < 0.01, * P < 0.05."

Fig. 4

Subcellular localization of FAB2 and FAD2"

Table 1

Agronomic trait investigation of germplasm resources"

花生品种
Peanut variety		 蛋白质含量
Protein content
(%)		 脂肪含量
Fatty acid content
(%)		 油酸
Oleic acid
(%)		 亚油酸
Linoleic acid
(%)		 硬脂酸
Stearic acid
(%)		 植物学类型
Botanical classification
冀甜 Jitian 24.10 42.04 30.28 44.87 4.28 珍珠豆型 Spanish type
汕油523 Shanyou 523 25.46 51.98 26.48 46.90 3.47 珍珠豆型 Spanish type
贺油11 Heyou 11 25.11 50.47 29.90 44.69 3.09 珍珠豆型 Spanish type
粤油18 Yueyou 18 26.03 51.24 22.04 51.34 3.62 珍珠豆型 Spanish type
徐花13 Xuhua 13 25.89 52.02 39.61 37.76 3.73 珍珠豆型 Spanish type
湛油12号 Zhanyou 12 20.98 55.68 33.59 42.77 2.39 珍珠豆型 Spanish type
花育30 Huayu 30 24.05 52.51 36.04 40.89 3.78 珍珠豆型 Spanish type
粤北种 Yuebeizhong 44.95 26.27 31.76 46.50 2.98 珍珠豆型 Spanish type
ICGV87123 25.73 48.86 29.46 44.89 2.32 珍珠豆型 Spanish type
粤油35 Yueyou 35 21.87 49.91 39.66 39.08 0.88 珍珠豆型 Spanish type
小粒种 Xiaolizhong 24.12 53.45 40.54 37.16 0.92 珍珠豆型 Spanish type
G40423 30.13 50.03 36.64 41.70 2.09 珍珠豆型 Spanish type
湛油1号 Zhanyou 1 29.88 46.76 28.99 43.87 3.74 珍珠豆型 Spanish type
闽花6号 Minhua 6 24.16 51.64 32.94 43.06 2.84 珍珠豆型 Spanish type
狮头企 Shitouqi 28.22 49.53 28.32 43.99 3.54 珍珠豆型 Spanish type
大明勾鼻 Daminggoubi 28.37 46.47 17.16 52.16 4.88 珍珠豆型 Spanish type
大罗大粒 Daluodali 46.42 23.75 30.61 47.88 2.22 普通型 Normal type
仲恺花1号 Zhongkaihua 1 22.81 52.10 31.10 45.22 1.95 珍珠豆型 Spanish type
宛花8908 Wanhua 8908 22.97 53.39 34.66 40.38 3.24 珍珠豆型 Spanish type
粤油3粒白 Yueyou 3 libai 25.45 48.53 41.38 37.20 0.94 珍珠豆型 Spanish type
文摘墩 Wenzhandun 28.08 48.88 47.66 34.59 0.30 珍珠豆型 Spanish type
ICGV92267 23.25 53.00 46.26 29.66 2.05 珍珠豆型 Spanish type
白沙1016 Baisha 1016 25.24 51.67 44.60 32.11 3.65 珍珠豆型 Spanish type
早花生1 Zaohuasheng 1 42.64 27.90 33.71 45.73 2.48 普通型 Normal type
西藏红 Xizanghong 26.01 51.18 34.15 41.31 2.50 珍珠豆型 Spanish type
花生品种
Peanut variety		 蛋白质含量
Protein content
(%)		 脂肪含量
Fatty acid content
(%)		 油酸
Oleic acid
(%)		 亚油酸
Linoleic acid
(%)		 硬脂酸
Stearic acid
(%)		 植物学类型
Botanical classification
狮选3号 Shixuan 3 30.32 47.70 19.26 51.07 4.26 珍珠豆型 Spanish type
HZ-152 24.11 40.90 15.49 53.25 5.12 珍珠豆型 Spanish type
蚂花2号 Mahua 2 27.09 51.04 31.05 43.64 2.62 珍珠豆型 Spanish type
汕油3号 Shanyou 3 45.69 25.08 35.41 45.52 1.78 珍珠豆型 Spanish type
仲恺花2号 Zhongkaihua 2 25.46 49.71 36.82 42.34 1.95 珍珠豆型 Spanish type
阳江铺地毡 Yangjiangpudizhan 43.61 25.65 37.56 42.33 2.57 珍珠豆型 Spanish type

Fig. 5

Phylogenetic of FAB2 protein in different peanut germplasm"

Fig. 6

Multiple alignment of FAB2 protein sequence in different peanut germplasm"

Table 2

Conserved elements in the promotor sequence of FAB2"

保守元件
Conversed elements		 结合序列
Binding sequence		 功能
Function
ABRE ACGTG Involved in the abscisic acid responsiveness
ARE AAACCA cis-acting regulatory element essential for the anaerobic induction
AT-rich sequence TAAAATACT Element for maximal elicitor-mediated activation
AT-TATA-box TATATA Transcription initiation
AuxRR-core GGTCCAT cis-acting regulatory element involved in auxin responsiveness
Box-4 ATTAAT Part of a conserved DNA module involved in light responsiveness
CAAT-box CAAT Common cis-acting element in promoter and enhancer regions
CGTCA-motif CGTCA cis-acting regulatory element involved in the MeJA-responsiveness
G-Box CACGTT cis-acting regulatory element involved in light responsiveness
GA-motif ATAGATAA Part of a light responsive element
GATA-motif AAGATAAGATT Part of a light responsive element
GC-motif CCCCCG Enhancer-like element involved in anoxic specific inducibility
GT1-motif GGTTAA Light responsive element
MRE AACCTAA MYB binding site involved in light responsiveness
MSA-like TCAAACGGT cis-acting element involved in cell cycle regulation
MYB TACCT MYB transcription factor binding site
MYC CATTG MYC transcription factor binding site
O2-site GATGATGTGG cis-acting regulatory element involved in zein metabolism regulation
P-box CCTTTTG Gibberellin-responsive element
TC-rich repeats ATTCTCTAAC cis-acting element involved in defense and stress responsiveness
TCCC-motif TCTCCCT Part of a light responsive element
TCT-motif TCTTAC Part of a light responsive element
TGA-element AACGAC Auxin-responsive element
TGACG-motif TGACG cis-acting regulatory element involved in the MeJA-responsiveness
WRE3 CCACT WRE transcription factor binding site
WUN-motif AAATTTCTT Wun transcription factor binding site
chs-CMA1a TTACTTAA Part of a light responsive element
as-1 TGACG As transcription factor binding site

Fig. 7

A proposed model of oleic acid regulating FAB2 expression A: schematic diagram of AT-rich region in FAB2 promotor sequence; B: a proposed putative model of oleic acid feedback regulating FAB2 expression."

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