Abstract
Polyacylated anthocyanins with multiple glycosyl and aromatic acyl groups tend to make flowers display bright and stable blue colours. However, there are few studies on the isolation and functional characterization of genes involved in the polyacylated anthocyanin biosynthesis mechanism, which limits the molecular breeding of truly blue flowers. Senecio cruentus is an important potted ornamental plant, and its blue flowers contain 3′,7-polyacylated delphinidin-type anthocyanins that are not reported in any other plants, suggesting that it harbours abundant gene resources for the molecular breeding of blue flowers. In this study, using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) analysis of blue, carmine and white colours of cineraria cultivars “Venezia” (named VeB, VeC, and VeW, respectively), we found that 3′,7-polyacylated anthocyanin, cinerarin, was the main pigment component that determined the blue colour of ray florets of cineraria. Based on the transcriptome sequencing and differential gene expression (DEG) analysis combined with RT- and qRT-PCR, we found two genes encoding uridine diphosphate glycosyltransferase, named ScUGT1 and ScUGT4; two genes encoding acyl-glucoside-dependent glucosyltransferases which belong to glycoside hydrolase family 1 (GH1), named ScAGGT11 and ScAGGT12; one gene encoding serine carboxypeptidase-like acyltransferase ScSCPL2; and two MYB transcriptional factor genes ScMYB2 and ScMYB4, that were specifically highly expressed in the ray florets of VeB, which indicated that these genes may be involved in cinerarin biosynthesis. The function of ScSCPL2 was analysed by virus-induced gene silencing (VIGS) in cineraria leaves combined with HPLC-MS/MS. ScSCPL2 mainly participated in the 3′ and 7-position acylation of cinerarin. These results will provide new insight into the molecular basis of the polyacylated anthocyanin biosynthesis mechanism in higher plants and are of great significance for blue flower molecular breeding of ornamental plants.
Highlights
Flower colour is one of the most important traits of ornamental plants
In this study, using comparative transcriptome analysis combined with gene expression, we found that ScUGT1 and ScUGT4 were highly expressed in the ray florets of VeB but almost not expressed in the ray florets of VeW and VeC (Figures 6D,E)
We found that ScUGT1 was clustered with DgpHBAGT, which is involved in the synthesis of p-hydroxybenzoyl-glucose, which acts as a zwitter donor in acylation and glucosylation in D. grandiflorum (Figure 6A) (Nishizaki et al, 2014)
Summary
Flower colour is one of the most important traits of ornamental plants. Blue floral colour primarily arises from anthocyaninss, which are modified by aromatic or fatty acyl groups that are commonly linked to the hydroxy groups of the glycosyl moieties. Anthocyanins modified with multiple aromatic acyl groups are often referred to as polyacylated anthocyanins, and the colour shifts to blue gradually as the number of acyl groups increases (Yoshida et al, 2009; Tanaka and Brugliera, 2013). Polyacylation at the 3 or 7-position of anthocyanin is more important for stable blue coloration, while the 3- or 5-position appears to provide only a reddish-purple colour (Matsuba et al, 2010; Nishizaki et al, 2013; Yoshida and Negishi, 2013). Many different polyacylated anthocyanin structures have been identified from different species, isolation and functional studies of genes involved in polyacylation modification are few, which limits the molecular breeding of truly blue flowers
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