Evidence for a NOD2-independent susceptibility locus for inflammatory bowel disease on chromosome 16p

Abstract

Heritable predisposition to inflammatory bowel disease (IBD) has been demonstrated by epidemiological and genetic analysis. Linkage of IBD to broad regions of chromosome 16 has been established by analysis of multiple populations. NOD2, located on proximal 16q, was recently identified as an IBD gene. As the linkage regions on chromosome 16 are large, we have investigated the possibility that NOD2 is not the only IBD gene located on this chromosome. A high-density experiment using 39 microsatellite markers was performed to identify additional regions of association, and to indicate areas of interest for further investigation. A triple-peaked configuration of the linkage curve with peak logarithm of odds (lod) scores of 2.7, 3.2, and 3.1 was observed on proximal 16p, proximal 16q, and central 16q, respectively. The cohort was stratified by coding individuals carrying the NOD2 single nucleotide polymorphism (SNP)8 and SNP13 "unknown." Significance at the central peak, corresponding to the genomic location of NOD2, then decreased from 3.2 to 1.2. The maximal lod scores on the proximal p-arm (lod = 2.1) and central q-arm (lod = 2.6) changed only moderately. An exploratory association analysis (TRANSMIT) yielded a strong lead at D16S3068 (P = 0.00028). The region around this marker was further investigated by using anonymous SNPs. An associated haplotype containing three SNPs was identified (peak significance P = 0.00027, IBD phenotype). On stratification based on NOD2 genotype, this significance increased to P = 0.0001. These results confirm the importance of NOD2 and provide evidence for a second IBD gene located on chromosome 16p.

Figure 1

(Top) The approximate locations of the linkage peaks obtained in other positive studies on chromosome 16 (six of the studies have been cited in the text; Ohmen et al. is ref. ; Parkes et al. is ref. 40). The respective lod scores or P values are given adjacent to the dark bar that indicates a peak location. The gray bars show the approximate location of the intervals with (lod_max = −1). An analysis of chromosome 16 microsatellite linkage to CD (Middle) and IBD (Bottom) is presented. The thick solid line represents the unstratified analysis. The thick gray line and the dashed line represent linkage analyses after the phenotypes of all carriers of the NOD2/SNP13 and NOD2/SNP8 and SNP13 mutation were set to “unknown,” respectively. ▵, The position of the NOD2 gene; ▾, the position of the centromere. The three markers (D16S3145, D16S415, and D16S3106) with the highest MLSs in the CD analysis are marked with large filled diamonds; all other marker positions are marked with small filled diamonds; and ⋄ indicates the position of the locus D16S3068. Linkage analysis was performed by using mapmaker/sibs (as integrated in genehunter 2). cM, centimorgan.

Figure 2

Association analysis (transmission distortion as calculated by TRANSMIT) of markers in the vicinity of the association signal in the vicinity of D16S3068. (Upper) The unstratified analysis of the German part of the extended cohort (no significant results were obtained in the United Kingdom part of the cohort, which have thus been omitted for clarity). (Lower) The results of the analysis of the NOD2-stratfied genotypes. The key of phenotypes and populations is given in Upper. The marker names and the corresponding physical map using the local BAC clones are shown below the association graphs.