Characterization of the 9-cis-epoxycarotenoid dioxygenase gene family and the regulation of abscisic acid biosynthesis in avocado

Avocado (Persea americana Mill. cv Lula) is a climacteric fruit that exhibi ts a rise in ethylene as the fruit ripens. This rise in ethylene is followe d by an increase in abscisic acid (ABA), with the highest level occurring j ust after the peak in ethylene production. ABA is synthesized from the clea vage of carotenoid precursors. The cleavage of carotenoid precursors produc es xanthoxin, which can subsequently be converted into ABA via ABA-aldehyde . Indirect evidence indicates that the cleavage reaction, catalyzed by 9-ci s-epoxycarotenoid dioxygenase (NCED), is the regulatory step in ABA synthes is. Three genes encoding NCED cleavage-like enzymes were cloned from avocad o fruit. Two genes, PaNCED1 and PaNCED3, were strongly induced as the fruit ripened. The other gene, PaNCED2, was constitutively expressed during frui t ripening, as well as in leaves. This gene lacks a predicted chloroplast t ransit peptide. It is therefore unlikely to be involved in ABA biosynthesis . PaNCED1 was induced by water stress, but expression of PaNCED3 was not de tectable in dehydrated leaves. Recombinant PaNCED1 and PaNCED3 were capable of in vitro cleavage of 9-cis-xanthophylls into xanthoxin and C-25-apocaro tenoids, but PaNCED2 was not. Taken together, the results indicate that ABA biosynthesis in avocado is regulated at the level of carotenoid cleavage.