Exploring the diversity in Citrus fruit colouration to decipher the relationship between plastid ultrastructure and carotenoid composition.

Differentiation of new and characteristic plastid ultrastructures during ripening of citrus fruits in both peel and pulp appears to be strongly correlated with the content and complement of carotenoids. Most of the species of the Citrus genus display a wide range in fruit colouration due to differences in carotenoids; however, how this diversity is related and may contribute to plastid differentiation and ultrastructure is currently unknown. To that end, carotenoid profile and plastid ultrastructure were compared in peel and pulp of three sweet oranges: the ordinary orange-coloured Navel, rich in β,β-xanthophylls, the yellow Pinalate mutant with an elevated content of colourless carotenes and reduced β,β-xanthophylls, and the red-fleshed Cara Cara with high concentration of colourless carotenes and lycopene in the pulp; and two grapefruits: the white Marsh, with low carotenoid content, and the red Star Ruby, accumulating upstream carotenes and lycopene. The most remarkable differences in plastid ultrastructure among varieties were detected in the pulp at full colour, coinciding with major differences in carotenoid composition. Accumulation of lycopene in Cara Cara and Star Ruby pulp was associated with the presence of needle-like crystals in the plastids, while high content of upstream carotenes in Pinalate pulp was related to the development of a novel plastid type with numerous even and round vesicles. The presence of plastoglobuli was linked to phytoene and xanthophyll accumulation, suggesting these structures as the main sites for the accumulation of these pigments. Peel chromoplasts were richer in membranes compared to pulp chromoplasts, reflecting their different biogenesis. In summary, differences in carotenoid composition and accumulation of unusual carotenoids are mirrored by the development of diverse and novel chromoplast types, revealing the plasticity of these organelles to rearrange carotenoids inside different structures to allow massive accumulation and thus contributing to the chemical stability of the carotenoids.