The discovery of new coding alleles of human CYP26A1 which are potentially defective in the metabolism of all-trans retinoic acid and their assessment in a recombinant cDNA expression system

Retinoic acid (RA) is a critical regulator of gene expression during embryonic development and in the maintenance of adult epithelial tissues. Genetic polymorphisms of CYP26A1 could cause inter-individual variation in the metabolism of retinoic acid, thus altering signaling during embryonic development. A total of 13 single nucleotide polymorphisms (SNPs) were identified in CYP26A1 in 92 racially diverse individuals (24 Caucasians, 24 African-Americans, 24 Asians and 20 individuals of unknown racial origin). Three of the 13 SNPS produced coding changes: R173S, F186L and C358R. These alleles were termed CYP26A1*2 , CYP26A1*3 , and CYP26A1*4 , respectively, by the Human Cytochrome P450 (CYP) Allele Nomenclature Committee at http://www.cypalleles.ki.se/ . cDNA constructs for wild-type and mutant alleles of CYP26A1 were constructed in a pcDNA3.1 expression vector containing a FLAG tag at the C-terminal end, which was used to identify and quantitate the CYP26A1 allelic proteins when expressed in COS-1 cells. Wild type CYP26A1 protein metabolized all-trans -retinoic acid (at-RA) to 4-oxo-RA, 4-OH-RA and 18-OH-RA as well as water-soluble metabolites. CYP26A1.3 (F186L) and CYP26A1.4 (C358R) allelic proteins exhibited significantly lower metabolism (40-80%) of at-RA than wild-type CYP26A1.1 protein. This study identifies two CYP26A1 coding alleles, CYP26A1*3 and CYP26A1*4 , which are predicted to be defective in retinoic acid metabolism based on the metabolism of at-RA by the recombinant proteins. This is the first study to identify coding alleles of CYP26A1 . The in vitro characterization of the recombinant allelic proteins suggests the need for future clinical studies of genotype/phenotype relationships of CYP26A1 in embryonic development.