Human RPE65 cDNA Clone(NM_000329.2)

The visual cycle in the retinal pigment epithelium (RPE) is essential for regenerating 11-cis retinal for photoreceptor function, with RPE65 retinol isomerase serving as the central enzymatic component. The researchers investigated the regulation of RPE65 expression in ARPE-19 and HEK293F cell models, which exhibit lower RPE65 mRNA and protein levels compared with native RPE. Using a series of constructs containing RPE65 ORFs with varying promoters, codon optimizations, and 3' UTR lengths, along with media supplementation experiments (nicotinamide vs. pyruvate), they demonstrated that transcriptional and metabolic status are primary determinants of RPE65 expression, whereas translational modulation plays a minor role. Additional experiments indicated that feeding rod outer segments (ROS) downregulates RPE65 in nicotinamide-grown cells, suggesting that core RPE functions, such as the visual cycle and phagocytosis, can be independently regulated.

Figure 1. Immunoblot analysis of ARPE-19 and HEK293F cells transfected with RPE65 constructs revealed higher protein expression in cells grown with nicotinamide and in constructs containing the full 3' UTR. (Postnikova OA, et al., 2025)