Subfunctionalization of D27 Isomerase Genes in Saffron.

Chromoplasts and chloroplasts contain carotenoid pigments as all- trans - and cis -isomers, which function as accessory light-harvesting pigments, antioxidant and photoprotective agents, and precursors of signaling molecules and plant hormones. The carotenoid pathway involves the participation of different carotenoid isomerases. Among them, D27 is a β-carotene isomerase showing high specificity for the C9-C10 double bond catalyzing the interconversion of all- trans - into 9- cis -β-carotene, the precursor of strigolactones. We have identified one D27 ( CsD27-1 ) and two D27-like ( CsD27-2 and CsD27-3 ) genes in saffron, with CsD27-1 and CsD27-3 , clearly differing in their expression patterns; specifically, CsD27-1 was mainly expressed in the undeveloped stigma and roots, where it is induced by Rhizobium colonization. On the contrary, CsD27-2 and CsD27-3 were mainly expressed in leaves, with a preferential expression of CsD27-3 in this tissue. In vivo assays show that CsD27-1 catalyzes the isomerization of all- trans - to 9- cis -β-carotene, and could be involved in the isomerization of zeaxanthin, while CsD27-3 catalyzes the isomerization of all- trans - to cis -ζ-carotene and all- trans - to cis -neurosporene. Our data show that CsD27-1 and CsD27-3 enzymes are both involved in carotenoid isomerization, with CsD27-1 being specific to chromoplast/amyloplast-containing tissue, and CsD27-3 more specific to chloroplast-containing tissues. Additionally, we show that CsD27-1 is co-expressed with CCD7 and CCD8 mycorrhized roots, whereas CsD27-3 is expressed at higher levels than CRTISO and Z-ISO and showed circadian regulation in leaves. Overall, our data extend the knowledge about carotenoid isomerization and their implications in several physiological and ecological processes.