Mol Cell Biol 32:763 (2012)
Evans DS1, Calton MA2, Kim MJ3, Kwok PY4, Miljkovic I5, Harris T6, Koster A6, Liu Y7, Tranah GJ1, Ahituv N3, Hsueh WC2, Vaisse C2.
Common genetic variants 3' of MC4R within two large linkage disequilibrium (LD) blocks
spanning 288 kb have been associated with common and rare forms of obesity. This large
association region has not been refined and the relevant DNA segments within the association
region have not been identified. In this study, we investigated whether common variants in the
MC4R gene region were associated with adiposity-related traits in a biracial population-based
study. Single nucleotide polymorphisms (SNPs) in the MC4R region were genotyped with a custom
array and a genome-wide array and associations between SNPs and five adiposity-related traits
were determined using race-stratified linear regression. Previously reported associations
between lower BMI and the minor alleles of rs2229616/Val103Ile and rs52820871/Ile251Leu were
replicated in white female participants. Among white participants, rs11152221 in a proximal 3'
LD block (closer to MC4R) was significantly associated with multiple adiposity traits, but
SNPs in a distal 3' LD block (farther from MC4R) were not. In a case-control study of severe
obesity, rs11152221 was significantly associated. The association results directed our
follow-up studies to the proximal LD block downstream of MC4R. By considering nucleotide
conservation, the significance of association, and proximity to the MC4R gene, we identified a
candidate MC4R regulatory region. This candidate region was sequenced in 20 individuals from a
study of severe obesity in an attempt to identify additional variants, and the candidate
region was tested for enhancer activity using in vivo enhancer assays in zebrafish and mice.
Novel variants were not identified by sequencing and the candidate region did not drive
reporter gene expression in zebrafish or mice. The identification of a putative insulator in
this region could help to explain the challenges faced in this study and others to link SNPs
associated with adiposity to altered MC4R expression.