Abstract
Chalcone isomerase gene (CHI) is a key gene that regulates the formation of yellow traits in petals. To reveal transcriptional regulatory mechanisms of CHI gene in petals of Paeonia lactiflora, we investigated the CHI expression using qPCR, the pigment content by HPLC, and methylation levels using BSP+Miseq sequencing in ‘Huangjinlun’ variety during different developmental stages including flower-bud stage (S1), initiating bloom (S2), bloom stage (S3), and withering stage (S4). Results showed that the expression level of CHI gene at S2 stage was significantly higher than that at other stages (P<0.05), and at S4 stage was extremely significantly lower than other stages (P<0.01). Besides, total anthocyanin, anthoxanthin, and flavonoid contents in petals presented a similar trend with CHI expression during developmental stages. A total of 16 CpG sites varying methylation levels were detected in CHI gene core promoter region, of which the methylation levels at mC-4 and mC-16 sites were extremely significantly negatively correlated with CHI mRNA expression (P<0.01). mC-16 site is located in the binding region of C/EBPα transcription factor, suggesting that methylation at the mC-16 site may inhibit the binding of C/EBPα to CHI promoter DNA, thereby regulating the tissue-specific expression of CHI gene. Our study revealed the expression pattern of CHI gene in petal tissues of P. lactiflora at different developmental stages, which is related to promoter methylation. Moreover, the important transcription regulation element–C/EBPα was identified, providing theoretical reference for in-depth study on the function of CHI gene in P. lactiflora.
Highlights
Herbaceous Peony (Paeonia lactiflora Pall.) is a traditional Chinese flower, and its color quality directly influences the ornamental merit and commercial value
chalcone isomerase (CHI), as the second known flavonoid biosynthesis-related enzyme, is one of the key enzymes required for the biosynthesis of flavonoids
Studies have shown that the lack of CHI gene expression or activity could seriously affect the flavonoid biosynthesis pathway in many plants, leading to a significant decrease in anthocyanidin and flavonoid contents [6,18], whereas the overexpression of CHI gene could increase flavonoid content [19]
Summary
Herbaceous Peony (Paeonia lactiflora Pall.) is a traditional Chinese flower, and its color quality directly influences the ornamental merit and commercial value. There are the rich resources of P. lactiflora varieties with different colors including pink, red, and purple in China; only one variety ‘Huangjinlun’ has yellow flower. The cultivation of new P. lactiflora varieties with novelty colors such as yellow is currently an important project for ornamental plant breeders. In the flavonoid biosynthetic pathway, the formation of yellow pigments is related to chalcone isomerase (CHI) gene [1]. CHI is a very stable enzyme participating in the early stage of flavonoid biosynthesis, and greatly accelerating the intramolecular cyclization of chalcones to form the flavonones. The CHI gene plays a very important role in the development of yellow flowers. Previous studies revealed that the expression levels of CHI gene directly affected the accumulation of upstream yellow chalcone, the downstream colorless or yellowish anthocyanins and red anthocyanins, leading to the changes in colors or flavonoids.
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