Comprehensive analysis of metabolomics and transcriptomics reveals varied tepal pigmentation across Gloriosa varieties

Gloriosa L. possesses exceptional ornamental value, with its floral hues exhibiting a wide range of variations. In this study, we employed sophisticated colorimetry, Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS), and transcriptome sequencing to investigate the phenotypic expression of tepal colors, the composition of carotenoids and anthocyanins, and the differential gene expression in four Gloriosa varieties during their full bloom phase. Our findings revealed that the redness of the tepals, indicated by higher a^* values, increased with the intensity of the red hue, while lighter colors corresponded to higher L^* values. Metabolomic analysis identified 50 carotenoids and 60 anthocyanins. It was observed that carotenoids primarily influence the yellow and orange color of Gloriosa tepals, with β-carotene, lutein, and zeaxanthin being the predominant carotenoids. Anthocyanins serve as the principal coloring agents in the orange, red and purple tepals of Gloriosa. High levels and proportions of cyanidins and pelargonidins are key contributors to the formation of red and purple tepals, while high levels and proportions of peonidins also play a significant role in purple coloration. In contrast, the presence of high levels and proportions of pelargonidins alone is a crucial factor in the formation of orange tepals. Transcriptomic data unearthed 57 and 92 candidate differentially expressed genes (DEGs) belong to carotenoid and anthocyanin biosynthesis pathway, respectively, with PSY, PDS, DFR, and ANS genes considered as critical genes for the differential accumulation of pigments of Gloriosa tepals. Weighted gene co-expression network analysis (WGCNA) revealed significant co-expression patterns between 217 transcription regulatory factors (TFs) and 8 carotenoid biosynthesis genes, and between 194 TFs and 41 anthocyanin biosynthesis genes. qRT-PCR verified the expression patterns of four carotenoid biosynthesis-related genes, eight anthocyanin biosynthesis-related genes, and three transcription regulatory factors. It was found that Cluster-121969.6 (MYB) gene is specifically expressed in the tepals of the four varieties (compared to stems and leaves) and shows a high consistency with the trend of anthocyanin content changes. This research provides new insights into the mechanisms underlying the formation of diverse floral colors in Gloriosa tepals.