Received for publication June 21, 2006.
Revised August 23, 2006.
Accepted for publication August 24, 2006.

Functional Genetic Diversity in the High-Affinity Carnitine Transporter OCTN2 (SLC22A5)

Abstract

Systemic carnitine deficiency (SCD) is a rare autosomal recessive disease resulting from defects in the OCTN2 (SLC22A5) gene, which encodes the high-affinity plasma membrane carnitine transporter. Although OCTN2 is fairly well studied in its relationship with SCD, little is known about the carrier frequency of disease-causing alleles of OCTN2, or of more common functional polymorphisms in this gene. To address these issues, we screened for genetic variants in the OCTN2 coding region by direct sequencing of the exons and flanking intronic region of OCTN2 in a large sample (n=276) of ethnically diverse subjects. Additionally, we established lymphoblastoid cell lines from subjects homozygous for either allele of the previously-identified promoter region variant, -207G>C. We found eight amino acid sequence variants of OCTN2, of which three (Phe17Leu, Leu144Phe, and Pro549Ser) were polymorphic in at least one ethnic group. When assayed for functional activity by expression in HEK-293 cells, using as probes both the endogenous substrate (L-carnitine) and the organic cation tetraethylammonium (TEA), three variants showed functional differences from the reference OCTN2 (Phe17Leu, Tyr449Asp, Val481Phe, p<0.05). Further studies of the F17L polymorphism showed a reduced V_max for L-carnitine transport, to approximately 50% of the reference OCTN2. Confocal microscopy studies using an OCTN2-GFP fusion protein showed that Phe17Leu had distinct subcellular localization from the reference OCTN2, with diffuse cytoplasmic retention of Phe17Leu, in contrast to reference OCTN2 which localized specifically to the plasma membrane. Lymphoblasts from subjects homozygous for the -207G allele showed increased L-carnitine transport compared to the -207C/C homozygotes (p<0.05). This study suggests that although loss-of-function mutations in OCTN2 are likely to be rare, common variants of OCTN2 found in healthy populations may contribute to variation in the disposition of carnitine and some clinically used drugs.

Key words: Amino Acid, Pharmacogenomic analyses, Cholesterol metabolism/lipoproteins

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