AUTHOR=Dong Tianyu , Song Shanglin , Wang Ying , Yang Ruixue , Chen Peilei , Su Jiuchang , Ding Xinru , Liu Yongkang , Duan Hongying 

TITLE=Effects of 5-azaC on Iridoid Glycoside Accumulation and DNA Methylation in Rehmannia glutinosa

JOURNAL=Frontiers in Plant Science

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/plant-science/articles/10.3389/fpls.2022.913717

DOI=10.3389/fpls.2022.913717

ISSN=1664-462X

ABSTRACT=<p>Iridoid glycoside is the important secondary metabolite and the main active component in <italic>Rehmannia glutinosa</italic>. However, the mechanisms that underlie the regulation of iridoid glycoside biosynthesis remain poorly understood in <italic>R. glutinosa</italic>. Herein, the analysis of RNA-seq data revealed that 3,394 unigenes related to the biosynthesis of secondary metabolites were identified in <italic>R. glutinosa</italic>. A total of 357 unigenes were involved in iridoid glycoside synthesis, in which the highly conservative genes, such as <italic>DXS, DXR, GPPS, G10H</italic>, and <italic>10HGO</italic>, in organisms were overexpressed. The analysis of the above genes confirmed that the co-occurrence ratio of <italic>DXS, DXR</italic>, and <italic>GPPS</italic> was high in plants. Further, our results showed that under normal and 5-azacytidine (5-azaC) treatment, the expression levels of <italic>DXS, DXR, GPPS, G10H</italic>, and <italic>10HGO</italic> were consistent with the iridoid glycoside accumulation in <italic>R. glutinosa</italic>, in which the application of the different concentrations of 5-azaC, especially 50 μM 5-azaC, could significantly upregulate the expression of five genes above and iridoid glycoside content. In addition, the changes in the spatiotemporal specificity of degree and levels of DNA methylation were observed in <italic>R. glutinosa</italic>, in which the hemi-methylation was the main reason for the change in DNA methylation levels. Similar to the changes in 5-methyl cytosine (5mC) content, the DNA demethylation could be induced by 5-azaC and responded in a dose-dependent manner to 15, 50, and 100 μM 5-azaC. Taken together, the expression of iridoid glycoside synthesis gene was upregulated by the demethylation in <italic>R. glutinosa</italic>, followed by triggering the iridoid glycoside accumulation. These findings not only identify the key genes of iridoid glycoside synthesis from <italic>R. glutinosa</italic>, but also expand our current knowledge of the function of methylation in iridoid glycoside accumulation.</p>