title: In Vitro Functional Analyses of Infrequent Nucleotide Variants in the Lactase Enhancer Reveal Different Molecular Routes to Increased Lactase Promoter Activity and Lactase Persistence.
abstract: The genetic trait that allows intestinal lactase to persist into adulthood in some 35% of humans worldwide operates at the level of transcription, the effect being caused by cis-acting nucleotide changes upstream of the lactase gene (LCT). A single nucleotide substitution, -13910 C>T, the first causal variant to be identified, accounts for lactase persistence over most of Europe. Located in a region shown to have enhancer function in vitro, it causes increased activity of the LCT promoter in Caco-2 cells, and altered transcription factor binding. Three other variants in close proximity, -13907 C>G, -13915 T>C and -14010 G>C, were later shown to behave in a similar manner. Here, we study four further candidate functional variants. Two, -14009 T>G and -14011 C>T, adjacent to the well-studied -14010 G>C variant, also have a clear effect on promoter activity upregulation as assessed by transfection assays, but notably are involved in different molecular interactions. The results for the two other variants (-14028 T>C, -13779 G>C) were suggestive of function, -14028*C showing a clear change in transcription factor binding, but no obvious effect in transfections, while -13779*G showed greater effect in transfections but less on transcription factor binding. Each of the four variants arose on independent haplotypic backgrounds with different geographic distribution.
authors.affiliation: Research Department of Genetics Evolution and Environment, University College London, London, UK.
authors.forename: Anke
authors.initials: A
authors.lastname: Liebert
authors.affiliation: Research Department of Genetics Evolution and Environment, University College London, London, UK.
authors.forename: Bryony L
authors.initials: BL
authors.lastname: Jones
authors.affiliation: Department of Science and Environment, Roskilde University, Roskilde, Denmark.
authors.forename: Erik Thomas
authors.initials: ET
authors.lastname: Danielsen
authors.affiliation: Department of Science and Environment, Roskilde University, Roskilde, Denmark.
authors.forename: Anders Krüger
authors.initials: AK
authors.lastname: Olsen
authors.affiliation: Research Department of Genetics Evolution and Environment, University College London, London, UK.
authors.forename: Dallas M
authors.initials: DM
authors.lastname: Swallow
authors.affiliation: Department of Science and Environment, Roskilde University, Roskilde, Denmark.
authors.forename: Jesper T
authors.initials: JT
authors.lastname: Troelsen
mesh_terms: D018938:Caco-2 Cells; D004742:Enhancer Elements, Genetic; D015870:Gene Expression; D015971:Gene Expression Regulation, Enzymologic; D056726:Genetic Association Studies; D006239:Haplotypes; D006801:Humans; D043322:Lactase / Q000096:biosynthesis / Q000235:genetics*; D007787:Lactose Intolerance / Q000235:genetics; D020641:Polymorphism, Single Nucleotide; D011401:Promoter Regions, Genetic
chemical_list: D043322:Lactase
keywords: Lactase persistence; enhancer; gel shift; supershift; transcription factor; transfection