Common variants in the 5' region of the leptin gene are associated with body mass index in men from the National Heart, Lung, and Blood Institute Family Heart Study

Abstract

Linkage of body mass index (BMI) to a broad region of chromosome 7q22-35 has been reported in multiple studies. We previously published a multipoint LOD score of 4.9 at D7S1804 for BMI from the National Heart, Lung, and Blood Institute Family Heart Study. Leptin (LEP), the human homolog of the mouse obesity (ob) gene, is positioned near the linkage peak and is the most prominent candidate gene in this region. Interest in LEP as a susceptibility gene for human obesity has led to numerous linkage and association studies, but the results of these studies are still controversial. In the present study, we employed family-based tests of association with both a quantitative measure of BMI adjusted for age and sex and a dichotomously defined obesity trait. We genotyped 29 single-nucleotide polymorphisms (SNPs) spanning 240 kb around the LEP gene in the 82 extended pedigrees with the strongest evidence for linkage. When the programs TRANSMIT and FBAT were used, a number of SNPs showed association in men but not women, for both the quantitative and qualitative trait definitions (P<.05). Five SNPs (H1328084, H1328083, H1328082, H1328081, and H1328080) positioned 2 kb beyond the previously defined promoter region showed strong association in single-marker and multiple-marker haplotype analysis. This five-marker haplotype (frequency 49% in this sample) is overtransmitted to obese offspring (P=.00005). All five of these SNPs are predicted to modify transcription-factor binding sites. This may indicate new functional variants in an extended promoter region of LEP.

Figures

Figure 1

LD block structure around LEP. Haplotype block structure, as depicted by Haploview, is shown. The five-color scheme (white to red) represents the increasing strength of LD. Values for D′ (×100) are shown, but those boxes with D′=1 are shaded in bright red and are empty. Cells with D′<1 are shades of pink or red. Blue represents D′=1 but with a low confidence estimate for D′.

Figure 2

Haplotype structure and diversity. Haplotype blocks and their frequencies were estimated using an accelerated EM algorithm implemented in Haploview. SNP numbers corresponding to figure 1 are listed above each column of alleles, and “▾” denotes the tag SNPs that designate a parsimonious haplotype for each block. Recombination rates from one block to the next are defined by a multiallelic value of D′. Haplotypes in adjacent blocks are connected by a thick line if they occur together with a frequency >10% and by a thin line if they occur together with a frequency >1%. For each SNP, “1” represents the common allele, and “2” represents the rare allele.

Figure 3

Haplotype analyses using one trio in each pedigree for a three-marker sliding window. Below each triad is the P value for that haplotype, the haplotype frequency, the direction of the transmitted haplotype (either overtransmitted [“O”] or undertransmitted [“U”]), and the global P value (median value of 20 runs), derived by evaluation of the transmissions of all haplotypes simultaneously. SNPs within the LEP gene are in boldface and are offset, and the approximate location within the LEP gene is shown below the haplotype frequencies. 1 = common allele; 2 = rare allele; InterDis = marker-to-marker distance (in bp); na = no haplotype resulted in P<.05; ns = not significant; Global-1nomf (P value) = global P value in the “one trio per pedigree” analysis. Celera SNP IDs are abbreviated to the final four digits. P values significant after FDR adjustment are indicated with an asterisk (*).

Figure 4

Genomic view of the LEP 5′ region, its polymorphisms, and the best five-marker haplotype. A, Genomic view of LEP mRNA and SNPs surrounding LEP (UCSC Genome Browser Web site). B, Polymorphisms at the 5′ region of LEP, with single-marker P values from TRANSMIT analysis and their respective putative TFBS, with the polymorphic site highlighted in red. No TFBS changes were found for G2548A and G1387A. na = no TFBS site identified for a specific short sequence.