Cloning And Functional Analysis Of BZIP Transcription Factor AaABF3 In Artemisia Annua L.

Artemisia annua L.is a traditional medicinal plant originating in China.Artemisinin is a sesquiterpene lactone that specifically exists in glandular trichomes of A.annua.Artemisinin-based combination therapy has been recognized as the best treatment for malaria by World Health Organization.However,the artemisinin contents in wild A.annua are very low,accounting for only 0.1%-1% of dry weight of leaves.Early studies have shown that plant hormone abscisic acid(ABA)plays an important role in increasing artemisinin content,but how ABA regulates artemisinin biosynthesis remains unclear.BZIP(Basic Leucin Zipper)transcription factors are involved in the transcriptional regulation of ABA-dependent gene expression pathway.In this study a bZIP transcription factor AaABF3 was characterized from A.annua and the molecular mechanism of ABA regulating artemisinin synthesis pathway was studied.The main results are as follows:1.The transcription factor AaABF3 was cloned from A.annua.Bioinformatics analysis showed that the gene was 1149 bp in length and encoded 382 amino acids.The expression pattern of AaABF3 was analyzed by GUS staining and qRT-PCR.It was found that AaABF3 was predominantly expressed in glandular trichomes of A.annua and was induced by ABA.It was also found that AaABF3 was located in the nucleus by subcellular localization analysis of AaABF3 protein.2.Dual-luciferase assay demonstrated that AaABF3 activated the expression of artemisinin synthesis gene ALDH1.Yeast one hybrid assay and electrophoretic mobility shift assay(EMSA)confirmed that AaABF3 could directly bind to the promoter of ALDH1,thus participating in regulating artemisinin biosynthesis.3.The over-expressing and RNA interference AaABF3 transgenic A.annua plants were obtained.High performance liquid chromatography assay(HPLC)showed that AaABF3 could promote the biosynthesis of artemisinin.It was verified that ABA regulated the biosynthesis of artemisinin through AaABF3 by detecting the response of AaABF3 and ALDH1 to ABA in transgenic plants.Through drought treatment of transgenic plants,it was speculated that AaABF3 may participate in drought resistance regulation of A.annua.