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Identification and characterization of a gene encoding a putative lysophosphatidyl acyltransferase from Arachis hypogaea

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Abstract

Lysophosphatidyl acyltransferase (LPAT) is the important enzyme responsible for the acylation of lysophosphatidic acid (LPA), leading to the generation of phosphatidic acid (PA) in plant. Its encoding gene is an essential candidate for oil crops to improve oil composition and increase seed oil content through genetic engineering. In this study, a full-length AhLPAT4 gene was isolated via cDNA library screening and rapid amplification of cDNA ends (RACE); our data demonstrated that AhLPAT4 had 1631 nucleotides, encoding a putative 43.8 kDa protein with 383 amino acid residues. The deduced protein included a conserved acyltransferase domain and four motifs (I–IV) with putative LPA and acyl-CoA catalytic and binding sites. Bioinformatic analysis indicated that AhLPAT4 contained four transmembrane domains (TMDs), localized to the endoplasmic reticulum (ER) membrane; detailed analysis indicated that motif I and motifs II–III in AhLPAT4 were separated by the third TMD, which located on cytosolic and ER luminal side respectively, and hydrophobic residues on the surface of AhLPAT4 protein fold to form a hydrophobic tunnel to accommodate the acyl chain. Subcellular localization analysis confirmed that AhLPAT4 was a cytoplasm protein. Phylogenetic analysis revealed that AhLPAT4 had a high homology (63.7–78.3%) with putative LPAT4 proteins from Glycine max, Arabidopsis thaliana and Ricinus communis. AhLPAT4 was ubiquitously expressed in diverse tissues except in flower, which is almost undetectable. The expression analysis in different developmental stages in peanut seeds indicated that AhLPAT4 did not coincide with oil accumulation.

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Abbreviations

CTAB:

hexadecyltrimethylammonium bromide

DAF:

days after flowering

ER:

endoplasmic reticulum

G3P:

sn-glycerol-3-phosphate

GFP:

green fluorescent protein

GPAT:

glycerol-3-phosphate acyltransferase

LPA:

lysophosphatidic acid

LPAT:

lysophosphatidyl acyltransferase

ORF:

open reading frame

PA:

phosphatidic acid

RACE:

rapid amplification of cDNA ends

TAG:

triacylglycerol

TMD:

transmembrane domain

UTR:

untranslated region

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Acknowledgements

This study was supported by the National Natural Science Foundation of China (31201239, 31071456), the Natural Science Foundation of Hebei Province, China (C2010001594), a grant from Modern Agro-industry Technology Research System (nycytx–19) and the Key Basic Research Program of Hebei Province Applied Basic Research Plan (10960122D). We gratefully thank Professor Shengyi Liu (OCRI, CAAS) and his laboratory staff for provision of plasmid pEGFP and technical assistance.

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Authors and Affiliations

  1. Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan, 430062, China

    Si-Long Chen, Jia-Quan Huang, Yong Lei, Yue-Ting Zhang, Xiao-Ping Ren, Yu-Ning Chen, Hui-Fang Jiang, Li-Ying Yan & Bo-Shou Liao

  2. Institute of Food and Oil Crops, Hebei Academy of Agriculture and Forestry Sciences, Laboratory of Crop Genetics and Breeding of Hebei Province, Shijiazhuang, 050031, China

    Si-Long Chen & Yu-Rong Li

  3. Graduate School of Chinese Academy of Agriculture Sciences, Beijing, 100081, China

    Si-Long Chen

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  1. Si-Long Chen
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Correspondence to Bo-Shou Liao.

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Corresponding editor: Indranil Dasgupta

Si-Long Chen and Jia-Quan Huang contributed equally to this work.

[Chen S-L, Huang J-Q, Lei Y, Zhang Y-T, Ren X-P, Chen Y-N, Jiang H-F, Yan L-Y, Li Y-R and Liao B-S 2012 Identification and characterization of a gene encoding a putative lysophosphatidyl acyltransferase from Arachis hypogaea. J. Biosci. 37 1–11] DOI 10.1007/s12038-012-9277-4

Supplementary materials pertaining to this article are available on the Journal of Biosciences Website at http://www.ias.ac.in/jbiosci/dec2012/supp/Chen.pdf

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Chen, SL., Huang, JQ., Lei, Y. et al. Identification and characterization of a gene encoding a putative lysophosphatidyl acyltransferase from Arachis hypogaea . J Biosci 37 (Suppl 1), 1029–1039 (2012). https://doi.org/10.1007/s12038-012-9277-4

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