Transcriptomic and metabolomic analyses reveal the mechanism of anthocyanin metabolism in H18 pepper leaves and the function of CaDFR1

1. The accumulation of anthocyanins on H18 pepper plants leads to a purple color in their leaves, with anthocyanins on the leaves decreasing as they grow and develop. 2. Anthocyanins produced with DHM as substrate occupy the main component of anthocyanins in H18 pepper leaves. 3. CaDRF1 plays an important role in the anthocyanin synthesis pathway of pepper leaves. Anthocyanins play a crucial role in plant growth, development, reproduction, and stress response. Additionally, anthocyanins enhance the quality of fruits and vegetables due to their antioxidant properties. While numerous previous research has been conducted on anthocyanins, limited information exists regarding their composition and the role of the anthocyanin pathway gene DFR (dihydroflavonol 4-reductase) in chili pepper leaves. In this study, we used a purple leaf pepper cultivar H18 with anthocyanins on the leaves decreasing as they grow and develop. Targeted anthocyanin metabolite assays revealed that the contents of delphinidin, malvidin, and petunidin derivatives followed the same trend as the overall anthocyanin content, with delphinidin derivatives being the predominant component of H18 pepper leaves. Transcriptome sequencing was performed on H18 leaves at four different stages. The results showed that DEGs at various stages were primarily associated with biological processes and flavonoid metabolic pathways. Through evolutionary tree and expression analysis, three candidate genes involved in DFR function were identified. Substrate catalysis assays of CaDFRs demonstrated that only CaDFR1 was active, catalyzing DHQ, DHM, and DHK. VIGS-mediated silencing of CaDFR1 resullted in a significant decrease reduction in anthocyanin levels in H18 pepper leaves and stems and along with a decreased reduction in the expression levels of other candidate functional genes in the anthocyanin metabolic pathway. This study identifies the key anthocyanin components in the leaves of H18 peppers and validates the function of CaDFR1, providing a theoretical foundation for modifying anthocyanin content in pepper plants through molecular breeding.