The effect on transcription efficiency of the apolipoprotein AI gene of DNA variants at the 5' untranslated region.

Abstract

Elevated circulating levels of high-density lipoprotein and apolipoprotein AI are associated with reduced coronary artery disease risk. We have shown that a C to T substitution at +83 bp and a G to A substitution at -75 bp of the apolipoprotein AI gene are both related to increased high-density lipoprotein levels in a healthy population but not in a coronary population, among whom the same mutations are associated with increased disease severity. In the present study, we explored the effects of these base changes on transcriptional efficiency in vitro. We directionally cloned (using polymerase chain reaction) the 5' region of the apolipoprotein AI gene (-281 to +330 bp) with GC, GT, and AC haplotypes into a pGL3-luciferase reporter gene basic vector, and transfected the constructed vectors into HepG2 cells. The cells carrying the T allele at the +83 bp site (GT 112.3 +/- 12.4) had the same transcriptional efficiency as those bearing the C allele (GC 126.3 +/- 9.6). However, for cells with the A allele at -75 bp there was a twofold decrease in transcription (AC 63.1 +/- 9.3) accompanied by similar changes in Luc+ mRNA levels; this reduced transcription was only present if the apolipoprotein AI leader sequence was included in the insert. While the findings are inconsistent with the T or A allele being associated with higher high-density lipoprotein levels, they are consistent with the finding that the alleles are associated with an increased coronary artery disease risk, and demonstrate that the 5' leader region of the apolipoprotein AI gene participates in regulating apolipoprotein AI transcription. They also suggest that other regions of the apolipoprotein AI gene may have an active role in such regulation, and that environmental effects may influence allele-specific expression.