Abstract
UV-B-induced flavonoid biosynthesis is epigenetically regulated by site-specific demethylation of AaMYB1, AaMYC, and AaWRKY TF-binding sites inAaPAL1promoter-causing overexpression ofAaPALgene inArtemisia annua. The present study was undertaken to understand the epigenetic regulation of flavonoid biosynthesis under the influence of ultraviolet-B radiation using Artemisia annua L. as an experimental model. In-vitro propagated and acclimatized plantlets were treated with UV-B radiation (2.8Wm-2; 3h), which resulted in enhanced accumulation of total flavonoid and phenolics content as well as eleven individual flavonoids measured through HPLC-DAC. Expression of eight genes (phenylanaline ammonia lyase, cinnamate-4-hydroxylase, 4-coumarate: CoA ligase; chalcone synthase, chalcone isomerase, cinnamoyl reductase, flavonoid-3'-hydroxylase, and flavones synthase) from upstream and downstream flavonoid biosynthetic pathways was measured through RT-PCR and RT-Q-PCR and all were variably induced under UV-B irradiation. Among them, AaPAL1 transcript and its protein were most significantly upregulated. Global DNA methylation analysis revealed hypomethylation of genomic DNA in A. annua. Further epigenetic characterization of promoter region of AaPAL1 revealed cytosine demethylation at five sites, which in turn caused epigenetic activation of six transcription factor-binding sites including QELEMENT, EBOXBNNAPA/MYCCONSENSUSAT, MYBCORE, MYBCOREATCYCB1, and GCCCORE. MYB transcription factors are positive regulators of flavonoid biosynthesis. Epigenetic activation of transcription-enhancing cis-regulatory elements in AaPAL1 promoter and subsequent overexpression of AaMYB1 and AaMYC and AaWRKY transcription factors under UV-B irradiation may probably be the reason for higher AaPAL1 expression and hence greater biosynthesis of flavonoids in A. annua L. The present study is the first report that provides mechanistic evidence of epigenetic regulation of flavonoid biosynthesis under UV-B radiation in A. annua L.
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