Lycopene ε-cyclase mediated transition of α-carotene and β-carotene metabolic flow in carrot fleshy root.

The accumulation of carotenoids such as xanthophylls, lycopene, and carotenes, is responsible for the color of carrot fleshy roots. The potential role of the DcLCYE encoding a lycopene ε-cyclase associated with carrot root color was investigated using cultivars with orange and red roots. The expression of DcLCYE in red carrot varieties was significantly lower than that in orange carrots at the maturity stage. Furthermore, red carrots accumulated a larger amount of lycopene and lower level of α-carotene. Sequence comparison and prokaryotic expression analysis revealed that amino acid differences in red carrots did not affect the cyclization function of DcLCYE. Analysis of DcLCYE catalytic activity revealed that it mainly involved ε-carotene, while a side activity on α-carotene and γ-carotene was also observed. Comparative analysis of the promoter region sequences indicated that differences in the promoter region may affect the transcription of DcLCYE. CaMV35S promoter-driven overexpression of DcLCYE was observed in red carrot 'Benhongjinshi'. Lycopene in transgenic carrot roots was cyclized to accumulate higher levels of α-carotene and xanthophylls, while β-carotene content decreased significantly. The expression levels of other genes in the carotenoid pathway were simultaneously upregulated. Editing DcLCYE in orange carrot 'Kurodagosun' by CRISPR/Cas9 resulted in a decrease in the α-carotene and xanthophyll contents. The relative expression levels of DcPSY1, DcPSY2 and DcCHXE were sharply increased in DcLCYE-edited mutants. The results of this study provided insights on DcLCYE function in carrots, which could be the basis for applications for creating colorful carrot germplasms.