Functional assessment of tyrosinase variants identified in individuals with albinism is essential for unequivocal determination of genotype to phenotype correlation.

Background Oculocutaneous albinism type 1 (OCA1), caused by pathogenic variations in the tyrosinase gene (TYR), is the most frequent and severe form of hypopigmentary disorders worldwide. While OCA1A manifests as a complete loss of melanin pigment, patients with OCA1B show residual pigmentation of skin, hair and eyes. Limited experimental evidence suggests retention of TYR in the endoplasmic reticulum (ER) causes OCA1 pathogenesis. However, a comprehensive functional analysis of TYR missense variations and correlation with genotype is lacking. Objectives Functional characterization of non-synonymous tyrosinase variants in OCA1 patients reported in the Albinism database (http://albinismdb.med.umn.edu/), the SNP database (dbSNP; http://www.ncbi.nlm.nih.gov/SNP/) and published literature, and an attempt to correlate them with reported/predicted phenotypes. Methods Thirty-four reported missense variations of TYR were subcloned by site directed mutagenesis, and the dual enzyme activities of the variant proteins were compared with wild type (WT). The degree of ER retention was also checked for each of the variants through Endoglycosidase H (Endo H) digestion followed by immunoprecipitation and densitometric analysis. Results Functional studies revealed one reported OCA1A variation with nearly 100% enzyme activity, 10 OCA1B variants lacking any enzyme activity, 8 non-synonymous SNPs with 30-60% of enzyme activity, and 3 SNPs that completely lacked activity altogether; the Endo H assay also corroborated these results. Conclusions Loss of enzyme activity of TYR variants was completely in agreement with ER retention across all variants examined. The results of the assay clearly established that determination of biological activity of identified variants in OCA patients is essential for correlating identified suspect genotype with the obvious phenotype of the disease. This article is protected by copyright. All rights reserved.