Mutational analysis of the proopiomelanocortin gene in Caucasians with early onset obesity

Abstract

OBJECTIVE: Mutations in the human gene encoding the polyhormone peptide proopiomelanocortin (POMC) are associated with obesity in rare cases and the gene co-localizes with a reported quantitative trait loci (QTL) for variations in circulating leptin levels and fat mass on human chromosome 2p21. In this study we have used polymerase chain reaction (PCR) and single strand conformation polymorphism (SSCP) analysis, to test whether variations in the human POMC gene are associated with human obesity. DESIGN AND SUBJECTS: Primary mutational analysis was performed on the coding region of the POMC gene and 500 bp of the putative promoter region, by single strand conformational analysis and sequencing, in 56 subjects with juvenile onset obesity (body mass index (BMI) ! 31 kgam 2 at the draft board examination). The prevalence of two polymorphisms were further studied in 156 obese and 205 control subjects, and in a population based cohort of 380 extensively characterized young healthy subjects. RESULTS: We have identi®ed a total of six gene variants, ®ve were silent nucleotide substitutions (No51(promoter) g?c, No670(5 H UTR)g?a, No4512(codon6)c?t CysaCys, No7726(codon116)c?t LeuaLeu) of which one was prevalent (No8246(3 H UTR)c?t) and one variant changed an amino acid (No8086(codon236)g?c ArgaGln). The amino acid substitution was only seen in one subject. Comparing the prevalence of the frequent No8246 silent polymorphism, in an association study comprising 156 subjects with juvenile onset obesity and 205 randomly sampled control subjects (mean BMI 23.5 AE 4.7 kgam 2 ), did not show any relationship to obesity. Also, comparing the prevalence of a known 9 bp insertionadeletion variant in the coding region of the gene between obese and lean, showed no association to obesity. Furthermore, analyzing a population based cohort of 380 young healthy Caucasians for the prevalent 3 H UTR polymorphism as well as the 9 bp insertionadeletion variant did not show any association to deviations in body fat contents or fasting serum leptin concentrations. CONCLUSION: In conclusion, it is unlikely that variations in the coding region and the putative promoter of the POMC gene are a major cause of juvenile onset human obesity.Keywords: juvenile onset obesity; proopiomelanocortin gene; obesity genetics; body fat; leptin IntroductionHuman obesity is a complex disorder determined by both inheritance and environment. Family studies 1 and studies of twins, 2,3 as well as adoptees reared apart 4,5,6 show that from 30 ± 70% of the variation in adiposity is determined by genetic factors. However, little is known about the mode of inheritance and no gene variants responsible for the common form of human obesity has so far been identi®ed.A recent study by Krude et al 7 has demonstrated that early onset severe obesity, adrenal insuf®ciency and red hair pigmentation, in two patients, is caused by mutations in the human proopiomelanocortin (POMC) gene. Moreover, a study by Comuzzie et al 8 has shown evidence of linkage between variations in circulating...

Proopiomelanocortin gene variants are associated with serum leptin and body fat in a normal female population

“…We found that the other two SNPs, 51 G/C and 7965 C/T, previously reported at low frequency in obese subjects, 11,16 occurred at even lower frequency in our normal female subjects: 51 G/C at 1.0% and 7965 C/T at 0.4%. The inclusion of these two rare SNPs was aimed at exposing any relatively strong effect in potential regulatory regions (51 G/C in the promoter and 7965 C/T in the 3 0 UTR) by examining a large sample.…”

“…It has been suggested that any causative mutations are more likely to occur in regulatory sequences, because coding changes could affect more than one of the overlapping cleavage products and have far-reaching effects on other physiological processes. 11 The associations of BMI, weight and total fat with SNPs in regions flanking the POMC gene exposed in this powerful study suggest that control of POMC expression may be influential in the determination of body weight. …”

“…The striking association of the 8246 C/T SNP with serum leptin in normal Caucasian subjects discovered by Suviolahti et al 13 demands replication, as it had been observed in only 70 female subjects and others had been unable to establish associations in, respectively, 194 11 and 370 12 nonobese female European Caucasians. Here, we have investigated association of four POMC SNPs with anthropometric variables, body fat and serum leptin in 2758 normal weight Caucasian female twins (mean age 47.4712.5 years), selected from the St Thomas' UK Adult Twin Registry (Twins UK) on the basis of available fasting plasma leptin data.…”

“…So far, no association has been found with obesity in case -control studies 11 -13 and smallscale studies (less than 350 subjects) have mainly failed to detect an association with BMI, serum leptin or body composition in either obese or nonobese subjects. 11,12 However, recently Suviolahti et al 13 in lean, but not in obese subjects, have found an association of the C allele with higher serum leptin (P ¼ 0.003). In this mixed sample, the female subjects were the main contributors to significance (P ¼ 0.004) and the association was even more pronounced in lean female carriers of 8246 CC/RsaI (À/À) or ( þ /À) genotype combination (Po0.0005).…”

“…In the POMC 5 0 flanking region, we tested RsaI and a second rare SNP, sited 630 bp upstream of the transcription start site in the 5 0 UTR, GenBank Accession V01510 nt 51 G/ C (not on the NCBI database), found at a frequency of 1.9% in obese Caucasians. 11 In the 3 0 UTR, we tested 8246 C/T and another SNP 281 bp upstream, GenBank Accession V01510 nt 7965 C/T; dbSNP rs2071345, found at a frequency of 1.0% in obese Caucasians 16 but at 29.0% in lean Japanese 17 (named 7285 C/T on the early numbering system 10 ). Our powerful study offered several prospects: (1) replication of the strong association of 8246C/T with serum leptin in normal Caucasian women; (2) exposure of small effects of the common SNPs that may have remained undetected previously; (3) exposure of strong effects of rare alleles untested in a normal population and (4) a more rigorous test of the reported RsaI/8246C/T haplotype association with serum leptin in normal subjects.…”