Abstract
An extensive body of work has documented the antioxidant role of xanthophylls (lutein and zeaxanthin) in human health and specifically how they provide photoprotection in human vision. More recently, evidence is emerging for the transcriptional regulation of antioxidant response by lutein/lutein cleavage products, similar to the role of β-carotene cleavage products in the modulation of retinoic acid receptors. Supplementation with xanthophylls also provides additional benefits for the prevention of age-related macular degeneration (AMD) and attenuation of Alzheimer’s disease symptoms. Mammalian β-carotene oxygenase 2 (BCO2) asymmetrically cleaves xanthophylls as well as β-carotene in vitro. We recently demonstrated that mouse BCO2 (mBCO2) is a functionally palmitoylated enzyme and that it loses palmitoylation when cells are treated with β-carotene. The mouse enzyme is the easiest model to study mammalian BCO2 because it has only one isoform, unlike human BCO2 with several major isoforms with various properties. Here, we used the same acyl-RAC methodology and confocal microscopy to elucidate palmitoylation and localization status of mBCO2 in the presence of xanthophylls. We created large unilamellar vesicle-based nanocarriers for the successful delivery of xanthophylls into cells. We demonstrate here that, upon treatment with low micromolar concentration of lutein (0.15 µM), mBCO2 is depalmitoylated and shows partial nuclear localization (38.00 ± 0.04%), while treatment with zeaxanthin (0.45 µM) and violaxanthin (0.6 µM) induces depalmitoylation and protein translocation from mitochondria to a lesser degree (20.00 ± 0.01% and 35.00 ± 0.02%, respectively). Such a difference in the behavior of mBCO2 toward various xanthophylls and its translocation into the nucleus in the presence of various xanthophylls suggests a possible mechanism for transport of lutein/lutein cleavage products to the nucleus to affect transcriptional regulation.
Highlights
Most consideration of the antioxidant function of xanthophylls, polar hydroxy carotenoids, has focused on their chemical role in quenching excited triplet states of singlet oxygen by virtue of their extended conjugated bond systems
Rabbit polyclonal antibody 186 was custom made against the mouse BCO2 multiple antigenic peptides (MAP)-SKFLQSDTYKANSAG peptide and 7055 rabbit polyclonal antibody was produced by co-immunization of the two human BCO2 MAP-SHENLHQEDL
Large Unilamellar Vesicles (LUVs) were prepared by hydration of lipid with carotenoids and assisting limonene, followed by extrusion and extraction of limonene to yield unilamellar vesicles with a narrow size distribution, as confirmed by Small-Angle X-ray Scattering (SAXS) and Dynamic Light Scattering (DLS), respectively (Figure 1)
Summary
Most consideration of the antioxidant function of xanthophylls, polar hydroxy carotenoids, has focused on their chemical role in quenching excited triplet states of singlet oxygen by virtue of their extended conjugated bond systems. It is postulated that the close proximity of xanthophylls allows effective energy transfer from excited all-trans-retinal to xanthophyll and prevents singlet oxygen generation by this photosensitizer [11]. By this mechanism, the largest part of excess energy can be transferred from potentially harmful triplets of photosensitizers to xanthophylls and dissipated as heat. A portion of dietary lutein is converted to meso-zeaxanthin, a stereoisomer of zeaxanthin, presumably in retinal pigment epithelium (RPE) by RPE65 isomerase [12] This preference for zeaxanthin has been suggested to be due to a greater antioxidant capacity, possibly due to the longer system of conjugated double bonds and membrane-stabilizing function of zeaxanthin (and meso-zeaxanthin) compared to lutein [12,13]. Accumulation in the nucleus of mBCO2 loaded with xanthophylls could directly affect gene expression and may provide a mechanism whereby xanthophylls/xanthophyll cleavage products could elicit a transcriptional response to oxidant stress
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