Lactase persistence DNA variant enhances lactase promoter activity in vitro: functional role as a cis regulatory element

Lactase persistence is a heritable, autosomal dominant, condition that results in a sustained ability to digest the milk sugar lactose throughout adulthood. The majority of the world's human population experiences a decline in production of the digestive enzyme lactase-phlorizin hydrolase during maturation. However, individuals with lactase persistence continue to express high levels of the lactase gene into adulthood. Lactase persistence has been strongly correlated with single nucleotide genetic variants, C/T−13910 and G/A−22018, located 13.9 and 22 kb upstream from the lactase structural gene. We aimed to characterize a functional role for the polymorphisms in regulating lactase gene transcription. DNA in the region of the C/T−13910 or G/A−22018 human lactase variants was cloned upstream of the 3.0 kb rat lactase gene promoter in a luciferase reporter construct. Human intestinal Caco-2 cells were transfected with the lactase variant/promoter–reporter constructs and assayed for promoter activity. A 200 bp region surrounding the C−13910 variant, associated with lactase non-persistence, results in a 2.2-fold increase in lactase promoter activity. The T−13910 variant, associated with lactase persistence, results in an even greater 2.8-fold increase. The DNA sequence of the C/T−13910 variants differentially interacts with intestinal cell nuclear proteins on EMSAs. AP2 co-transfection results in a similar repression of the C/T−13910 variant/promoter–reporter constructs. The DNA region of the C/T−13910 lactase persistence/non-persistence variant functions in vitro as a cis element capable of enhancing differential transcriptional activation of the lactase promoter. Such differential regulation by the C and T variants is consistent with a causative role in the mechanism specifying the lactase persistence/non-persistence phenotypes in humans.