Investigation of a genome wide association signal for obesity: synthetic association and haplotype analyses at the melanocortin 4 receptor gene locus

André Scherag, Ivonne Jarick, Jessica Grothe, Heike Biebermann, Susann Scherag, Anna-Lena Volckmar, Carla Ivane Ganz Vogel, Brandon Greene, Johannes Hebebrand, Anke Hinney, André Scherag, Ivonne Jarick, Jessica Grothe, Heike Biebermann, Susann Scherag, Anna-Lena Volckmar, Carla Ivane Ganz Vogel, Brandon Greene, Johannes Hebebrand, Anke Hinney

Abstract

Background: Independent genome-wide association studies (GWAS) showed an obesogenic effect of two single nucleotide polymorphisms (SNP; rs12970134 and rs17782313) more than 150 kb downstream of the melanocortin 4 receptor gene (MC4R). It is unclear if the SNPs directly influence MC4R function or expression, or if the SNPs are on a haplotype that predisposes to obesity or includes functionally relevant genetic variation (synthetic association). As both exist, functionally relevant mutations and polymorphisms in the MC4R coding region and a robust association downstream of the gene, MC4R is an ideal model to explore synthetic association.

Methodology/principal findings: We analyzed a genomic region (364.9 kb) encompassing the MC4R in GWAS data of 424 obesity trios (extremely obese child/adolescent and both parents). SNP rs12970134 showed the lowest p-value (p = 0.004; relative risk for the obesity effect allele: 1.37); conditional analyses on this SNP revealed that 7 of 78 analyzed SNPs provided independent signals (p≤0.05). These 8 SNPs were used to derive two-marker haplotypes. The three best (according to p-value) haplotype combinations were chosen for confirmation in 363 independent obesity trios. The confirmed obesity effect haplotype includes SNPs 3' and 5' of the MC4R. Including MC4R coding variants in a joint model had almost no impact on the effect size estimators expected under synthetic association.

Conclusions/significance: A haplotype reaching from a region 5' of the MC4R to a region at least 150 kb from the 3' end of the gene showed a stronger association to obesity than single SNPs. Synthetic association analyses revealed that MC4R coding variants had almost no impact on the association signal. Carriers of the haplotype should be enriched for relevant mutations outside the MC4R coding region and could thus be used for re-sequencing approaches. Our data also underscore the problems underlying the identification of relevant mutations depicted by GWAS derived SNPs.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Genomic region encompassing the melanocortin…
Figure 1. Genomic region encompassing the melanocortin 4 receptor gene (MC4R; location indicated by the vertical bar).
Displayed are results of transmission disequilibrium tests of single markers and selected two-marker haplotypes in the detection sample of 424 obesity trios (two-sided exact p-values). The black dots indicate single marker TDT results for all 78 SNPs whereas the dots surrounded by a circle highlight the 8 SNPs used for two-marker haplotype construction (selected based on conditional analyses). The region covered by each two-marker haplotype combination is displayed as a line (only those haplotypes with a p-value≤0.1 and whose p-value was below the p-value of each single-marker test are displayed). Surrounding double circles indicate those SNPs (single-marker TDT) of the best supported haplotype result; Haplo 3 spans MC4R. In addition, grey lines indicate recombination rates according to HAPMAP CEU.

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