The variable number of tandem repeats element in DATI regulates in vitro dopamine transporter density -: art. no. 55

Background: A 40-bp variable number of tandem repeats (VNTR) polymorphism exists in the 15(th) exon of DATI, the gene encoding the human dopamine transporter (DAT). Though the VNTR resides in a region encoding the 3' untranslated region (UTR) and does not alter the protein's amino acid sequence, the prevalent 10-repeat variant has shown both linkage and association to Attention Deficit Hyperactivity Disorder (ADHD). In this study, we examined the effects of the DATI VNTR on measures of in vitro DAT expression and pharmacology. A series of four DATI constructs, each containing the DATI coding region, but varying with respect to the downstream presence or content of the 3' UTR, were engineered and stably transfected into an HEK-293 variant using Flp-In integration, an enzyme-mediated, site-specific recombination technology. Results: [H-3] Win 35,428 saturation binding assays and DAT immunoblots revealed statistically significant differences in DAT expression attributable to DATI genotype. Cells harboring the 10-repeat DATI variant were characterized by a B-max approximately 50% greater than cells with the 9-repeat VNTR; those containing only the DATI coding region or the coding region flanked by a truncated 3' UTR resulted in greater DAT density than either of the naturalistic 9- and 10-repeat variants. Competition binding assays showed no statistically significant DATI genotype effects on the DAT affinity for methylphenidate, a finding consistent with the positional location of the VNTR. Conclusion: This study identified the DATI VNTR as a functional polymorphism and provides an interpretive framework for its association with behavioral phenotypes.