Mutational screening of the CART gene in obese children - Identifying a mutation (Leu34Phe) associated with reduced resting energy expenditure and cosegregating with obesity phenotype in a large family

Cocaine- and amphetamine-regulated transcript (CART) inhibits feeding and i nduces the expression of c-Fos in hypothalamic areas implicated in appetite regulation. Furthermore, the CART peptide is found in neurons regulating s ympathetic outflow, which in turn play an integral role in regulating body temperature and energy expenditure. The CART gene was screened by single-st rand conformation polymorphism and automatic sequencing in 130 (72 girls) u nrelated obese Italian children and adolescents. Their Z-scores (mean +/- S D) of relative to BMI percentiles was 3.9 +/- 1.8, and the average age at o besity onset was 4.7 +/- 2.6 years. Two previously described silent polymor phisms were found in the 3 ' untranstated region: an adenine deletion at po sition 1457 in 9 patients (allele frequency 0.035) and an A/G substitution at position 1475 in 11 patients (allele frequency 0.042). We found no diffe rence between the obese patients heterozygous for one of these polymorphism s and those patients homozygous, for the wild allele with respect to their age of obesity onset, BMI Z-scores, and leptin levels. A missense mutation of G729C resulting in the substitution of Leu with Phe at codon 34, within the NH2-terminal CART region, has been detected in the heterozygous state i n a 10-year-old obese boy who has been obese since the age of 2 years. The patient belongs to a large family of obese subjects. The mutation cosegrega ted with the severe obesity phenotype over three generations and was not fo und in the control population. Resting metabolic rates were lower than expe cted in the propositus (-14%) and his mother (-16%), who carried the mutati on. Leucine at codon 34, conserved in this position in the human and in the rat sequences, immediately precedes a couple of lysine residues that may w ell represent a dibasic processing site. The Leu34Phe mutation might alter the susceptibility to proteolysis of this potential processing site, likely altering the CART effect on thermogenesis and energy expenditure.