DNA Methylation Of The Chalcone Ismerase Gene Promoter Region In Lithocarpus Polystachyus And Its Effects On Flavoniod Metablism

Lithocarpus polystachyus Rehd is a plant resource with three functions: medicine,food and tea,and has a history of application for thousands of years.Flavonoid metabolitese are its main active components.Metabolite biosynthesis is regulated by gene expression,which is altered by DNA methylation in the promoter region.Chalcone isomerase(CHI)gene encodes a key enzyme in flavonoid pathway,and the expression of L.polystachyus CHI(Lp CHI)is closely related to the synthesis of flavonoid metabolites.To analyze the DNA methylation site of the Lp CHI promoter and its effect on gene expression and metabolite accumulation.Lp CHI promoter sequence was cloned using thermal asymmetric interlaced PCR(TAIL-PCR).The Cp G island of the gene promoter was predicted using online software,and the DNA methylation sites of the Lp CHI promoter was investigated through bisulfite sequencing.The results show that the full-length promoter sequence of CHI was 2141 bp,with 32 types,135 cis-acting elements,3 Cp G islands.The 16 DNA methylation sites and the proportions of three types of Lp CHI promoter DNA methylation are 7.5%(Type A),68.75%(Type B),18.75%(Type C),determined by bisulfite sequencing.Transcriptome sequencing shows that Lp CHI is strongly up-regulated in Lp CHI promoter methylation Type A but down-regulated in Lp CHI promoter methylation Type B and Type C.The expression of Lp CHI is not significantly different between Type B and Type C.Besides,nine kinds of differentially expressed transcription factors bind to seven DNA methylation sites of the Lp CHI promoter region and regulate Lp CHI expression.The results of metabolomics show that differentially accumulated flavonoids in Lp CHI promoter methylation Type A are higher than in Lp CHI promoter methylation Type B and Type C.Additionally,we found the Lp CHI expression and flavonoids accumulation has positive correlations.These results show that the effect of DNA methylation site-specific on gene transcription is great than the effect of overall promoter DNA methylation on gene transcription and further reveal the mechanisms of flavonoid genes regulating metabolite accumulation.Figure 36;Table 26;Reference 89...