Antisense inhibition of the beta-carotene hydroxylase enzyme in Arabidopsis and the implications for carotenoid accumulation, photoprotection and antenna assembly.

The xanthophylls are oxygenated carotenoids and are important structural components of the photosynthetic apparatus. Xanthophylls contribute to the assembly and stability of light-harvesting complex apoproteins (LHC) and contribute to photoprotection via non-photochemical quenching of chlorophyll fluorescence (NPQ) in oxygenic photosynthetic organisms. Previously, mutations have been described that disrupt many steps in the xanthophyll biosynthetic pathway. However, there are no definitive reports of a lesion that effects the beta-hydroxylase enzyme, which catalyzes hydroxylation of the beta-rings of beta-carotene and alpha-carotene, and is thus necessary for synthesis of essentially all xanthophylls of higher plant chloroplasts. We have utilized an antisense approach to effectively reduce levels of beta-hydroxylase in Arabidopsis thaliana in order to examine how a reduction in this enzyme impacts carotenoid biosynthesis and plant viability. Expression of the antisense beta-hydroxylase transgene resulted in a maximal reduction in violaxanthin of 64% and a maximal reduction in neoxanthin of 41%. This reduction was reflected in a 22% increase in beta-carotene and a reduction in the total carotenoid pool, whereas lutein levels were relatively unaltered. Despite the reduction in violaxanthin and neoxanthin, the antisense beta-hydroxylase plants had a wild-type complement of chlorophylls and LHCs on a fresh weight basis. Under high light stress, the unconverted pool of violaxanthin was the same size as in wild type and thus there was an even greater proportional reduction in zeaxanthin of 75%. Despite this marked decrease in zeaxanthin, NPQ only declined by 16%.