Study on the Coloration Mechanism Underlying Leaf Variegation in Osmanthus Fragrans ‘Yuntian Caigui’

Colored-leaf plants, prized for their distinctive hues, are highly favored and have become indispensable in landscape design. Variegated varieties of Osmanthus fragrans are highly valued for their unique ornamental characteristics, which are attributed to differently colored patches on their leaves. However, the molecular mechanism of this character is still unclear. In this study, the yellow sectors of the variegated leaves contained significantly lower levels of chlorophyll a, chlorophyll b, and carotenoids compared to both the green sectors and the reverted green leaves. Transmission electron microscopy further demonstrated abnormal chloroplast development in the yellow sectors, characterized by proliferated stromal thylakoids and disorganized stacking of granum thylakoid. In contrast, the green sectors and reverted leaves displayed well-organized thylakoid structures. These structural anomalies resulted in impaired photosynthetic function, reduced electron transfer efficiency, and diminished performance of PSII reaction centers. Whole-genome resequencing analysis identified 36,564,922 single-nucleotide polymorphisms and 6,339,078 insertion-deletion sites, of which 2,545 showed different distribution. Notably, a specific AA-insertion was detected within the promoter region of the OfCOL4 gene. The expression level of OfCOL4 was significantly higher in the yellow sectors than in the green sectors of variegated leaves. Transient overexpression of OfCOL4 in Nicotiana benthamiana leaves led to a significant reduction in the levels of both chlorophyll and carotenoids, resulting in leaf bleaching. This study provides a valuable genetic resource and lays the groundwork for molecular breeding focused on developing novel ornamental plants with leaf variegation