Fine mapping of the celiac disease-associated LPP locus reveals a potential functional variant

Abstract

Using the Immunochip for genotyping, we identified 39 non-human leukocyte antigen (non-HLA) loci associated to celiac disease (CeD), an immune-mediated disease with a worldwide frequency of ∼1%. The most significant non-HLA signal mapped to the intronic region of 70 kb in the LPP gene. Our aim was to fine map and identify possible functional variants in the LPP locus. We performed a meta-analysis in a cohort of 25 169 individuals from six different populations previously genotyped using Immunochip. Imputation using data from the Genome of the Netherlands and 1000 Genomes projects, followed by meta-analysis, confirmed the strong association signal on the LPP locus (rs2030519, P = 1.79 × 10(-49)), without any novel associations. The conditional analysis on this top SNP-indicated association to a single common haplotype. By performing haplotype analyses in each population separately, as well as in a combined group of the four populations that reach the significant threshold after correction (P < 0.008), we narrowed down the CeD-associated region from 70 to 2.8 kb (P = 1.35 × 10(-44)). By intersecting regulatory data from the ENCODE project, we found a functional SNP, rs4686484 (P = 3.12 × 10(-49)), that maps to several B-cell enhancer elements and a highly conserved region. This SNP was also predicted to change the binding motif of the transcription factors IRF4, IRF11, Nkx2.7 and Nkx2.9, suggesting its role in transcriptional regulation. We later found significantly low levels of LPP mRNA in CeD biopsies compared with controls, thus our results suggest that rs4686484 is the functional variant in this locus, while LPP expression is decreased in CeD.

Figure 1.

Fine mapping of the LPP region. (A) Localization of the LPP gene on chromosome 3 and the LD region. (B) Meta-analysis of imputation and Immunochip (iCHIP) results. Open circles are SNPs genotyped by the iCHIP, black dots represent SNPs imputed by GoNL and gray dots represent SNPs imputed by the 1000 Genomes Project (1kG). (C) SNPs in the fine-mapping region overlapping to ENCODE regulatory regions, including DNase I hypersensitive sites, histone modifications determined by ChIP-seq and transcription factor-binding site ChIP signals. Signal peaks are depicted as darker regions. Evolutionary conserved sites are also shown. *14 SNPs that construct the core-haplotype. **7 SNPs in orange differentiate the risk from the non-risk haplotype. The functional candidate SNP rs4686484 is depicted in red. The top meta-analysis SNP is in blue. (D) Sequencing of the functional candidate SNP predicted to change the binding site of the shown transcription factors. Prot, protective; Ref, reference.