Light-induced Artemisinin Biosynthesis Is Regulated By AaHY5 Transcription Factor In Artemisia Annua

Artemisinin is a sesquiterpene lactone endoperoxide extracted from a traditional Chinese medicinal plant Artemisia annua.Artemisinin-based combination therapy(ACTs)is recommended as the best treatment of malaria by the World Health Organization(WHO).A.annua is the only plant source of artemisinin,but the content of artemisinin in A.annua is too low.Therefore,it is important to study the molecular regulation mechanism of artemisinin biosynthesis for the improvement of artemisinin content in A.annua.It was found that a variety of environmental signals and phytohormone signals can affect the content of artemisinin.Jasmonic acid(JA)and light signals can promote the biosynthesis of artemisinin.However,the molecular mechanism of light signal promoting artemisinin biosynthesis is not clear.Further studies have found that JA promotes the expression of artemisinin biosynthesis genes depending on light.However,the molecular mechanism of signal interaction has not been studied.In order to reveal the above scientific problems,following studies were carried out:1.Transcriptome analysis of genes associated with the artemisinin biosynthesis by jasmonic acid treatment under the light in Artemisia annuaIt was found that the increase of artemisinin biosynthesis by JA was dependent on light.However,the relationship between the two signal pathways mediated by JA and light remains unclear.In this study,the leaves of A.annua seedlings after 24 h continuous light(Light),24 h dark treatment(Dark),4 h MeJA treatment under the continuous light conditions(Light-MeJA-4h)and 4 h MeJA treatment under the dark conditions(Dark-MeJA-4h)were collected and the transcriptome sequencing using Illumina HiSeq 4000 System was performed.A total of 266.7 million clean data were produced which were subsequently assembled into 185,653 unigenes,with an average length of 537 bp.Among them,59,490 unigenes were annotated and classified based on the public information.Differential expression analyses were performed between Light and Dark,Light and Light-MeJA-4h,Dark and Dark-MeJA-4h,Light-MeJA-4h and Dark-MeJA-4h,respectively.1,588 transcription factors(TFs)were identified,which were divided into 55 TF families,with 284 TFs down-regulated in the Dark relative to Light and 96 TFs up-regulated in the Light-MeJA-4h relative to Light.Eight TFs were selected as candidates for regulating the artemisinin biosynthesis and one of them was validated to be involved in artemisinin transcriptional regulation by Dual-Luciferase(Dual-LUC)assay.The transcriptome data in this study offer a comprehensive transcriptional expression pattern influenced by the MeJA and light in A.annua seedling,which will serve as a valuable resource for further studies on transcriptional regulation mechanism underlying artemisinin biosynthesis.2.Light-induced artemisinin biosynthesis is regulated by the bZIP transcription factor AaHY5 in Artemisia annuaLight has been proposed to play an important role in the activation of artemisinin biosynthesis.In this study,the basic leucine zipper transcription factor AaHY5 was found to act as a key regulator of light-induced biosynthesis of artemisinin.The result showed that AaHY5 expression overlaped with those of artemisinin biosynthesis genes in response to light and in A.annua tissues.Analysis of AaHY5 over-expression and RNAi-suppression A.annua lines suggests that AaHY5 is a positive regulator of the expression of artemisinin biosynthesis genes and of artemisinin accumulation.It was showed that AaHY5 complemented the hy5 mutant in Arabidopsis thaliana.These data further suggests that AaHY5 interacts with AaCOP1,the ubiquitin E3 ligase CONSTITUTIVE PHOTOMORPHOGENIC1 in A.annua.The yeast one hybrid and transient expression assays demonstrate that AaHY5 acts via the transcription factor GLANDULAR TRICHOME-SPECIFIC WRKY 1(AaGSW1)in artemisinin regulation.In summary,this study presents a novel regulator of artemisinin gene expression and propose a model in which AaHY5 indirectly controls artemisinin production in response to the changing light conditions.