Molecular cloning, characterization, and promoter analysis of the isochorismate synthase (AaICS1) gene from Artemisia annua

Isochorismate synthase (ICS) is a crucial enzyme in the salicylic acid (SA) synthesis pathway. The full-length complementary DNA (cDNA) sequence of the ICS gene was isolated from Artemisia annua L. The gene, named AaICS1 , contained a 1710-bp open reading frame, which encoded a protein with 570 amin...

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Bibliographic Details
Published in:Journal of Zhejiang University. B. Science 2017-08, Vol.18 (8), p.662-673
Main Authors: Wang, Lu-yao, Zhang, Ying, Fu, Xue-qing, Zhang, Ting-ting, Ma, Jia-wei, Zhang, Li-da, Qian, Hong-mei, Tang, Ke-xuan, Li, Shan, Zhao, Jing-ya
Format: Article
Language:English
Subjects:
Amino acids
Artemisia annua
Artemisinin
Binding sites
Bioinformatics
Biomedical and Life Sciences
Biomedicine
Cloning
Comparative studies
Complementary DNA
Deoxyribonucleic acid
DNA
Drought
Gene expression
High performance liquid chromatography
Homology
Isochorismate synthase
Leaves
Liquid chromatography
Nucleotide sequence
Plant tissues
Polymerase chain reaction
Regulatory sequences
Ribonucleic acid
RNA
RNA-mediated interference
Salicylic acid
Transcription
Transgenic plants
Wounding
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Summary:Isochorismate synthase (ICS) is a crucial enzyme in the salicylic acid (SA) synthesis pathway. The full-length complementary DNA (cDNA) sequence of the ICS gene was isolated from Artemisia annua L. The gene, named AaICS1 , contained a 1710-bp open reading frame, which encoded a protein with 570 amino acids. Bioinformatics and comparative study revealed that the polypeptide protein of AaICS1 had high homology with ICS s from other plant species. Southern blot analysis suggested that AaICS1 might be a single-copy gene. Analysis of the 1470-bp promoter of AaICS1 identified distinct cis -acting regulatory elements, including TC-rich repeats, MYB binding site (MBS), and TCA-elements. An analysis of AaICS1 transcript levels in multifarious tissues of A. annua using quantitative real-time polymerase chain reaction (qRT-PCR) showed that old leaves had the highest transcription levels. AaICS1 was up-regulated under wounding, drought, salinity, and SA treatments. This was corroborated by the presence of the predicted cis -acting elements in the promoter region of AaICS1 . Overexpressing transgenic plants and RNA interference transgenic lines of AaICS1 were generated and their expression was compared. High-performance liquid chromatography (HPLC) results from leaf tissue of transgenic A. annua showed an increase in artemisinin content in the overexpressing plants. These results confirm that AaICS1 is involved in the isochorismate pathway.
ISSN:1673-1581
1862-1783
DOI:10.1631/jzus.B1600223