An upstream polymorphism associated with lactase persistence has increased enhancer activity

Abstract

Background & Aims : Intestinal lactase activity declines during childhood in some humans. This phenotypic polymorphism of lactase persistence or nonpersistence into adult life has been shown in a recent study to be 100% associated with a T/C nucleotide polymorphism at position −13910 and approximately 97% with an A/G nucleotide polymorphism at position −22018. The aim of this study was to investigate the role of these nucleotide polymorphisms for lactase-phlorizin hydrolase (LPH) gene expression. Methods : The −13910 and −22018 regions were cloned from lactase-persistent and -nonpersistent individuals, and the regions were analyzed for gene regulatory activity of a luciferase reporter gene by transfection experiments using the intestinal cell line Caco-2. Electrophoretic mobility shift assays (EMSAs) were used to investigate protein/DNA interactions with the −13910 sequence. Results : We show that the −13910 region contains a strong enhancer. The −13910 regions from both lactase persistent (−13910T variant) and lactase nonpersistent (−13910C variant) have enhancer activity. However, the −13910T variant enhances the LPH promoter approximately 4 times more than the −13910C variant when analyzed in differentiated Caco-2 cells. A nuclear factor from both an intestinal and a nonintestinal extract binds strongly to the −13910T variant whereas the binding to the −13910C variant is much weaker. Conclusions : The discovery of a functional difference between the 2 alleles at position −13910 supports the notion that the molecular difference between lactase persistence and nonpersistence is caused by the mutation at position −13910.