Functional Characterization Of A Novel Human Melanocortin 4 Receptor Mutation

Program: Abstracts - Orals, Featured Poster Presentations, and Posters
Session: SAT 697-707-Obesity Pathophysiology
Saturday, June 15, 2013: 1:45 PM-3:45 PM
Expo Halls ABC (Moscone Center)

Poster Board SAT-706
Patric JD Delhanty*1, Elise Bouw1, Martin Huisman1, Resie Vervenne2, Axel PN Themmen1, Aart Jan van der Lely1 and Erica LT van den Akker1
1Erasmus MC, Rotterdam, Netherlands, 2VUMC, Amsterdam, Netherlands
Mutations in the MC4R gene are the most common cause of monogenic obesity. The melanocortin 4 receptor (MC4R) is expressed in the hypothalamus and is essential for regulation of appetite and energy expenditure. MC4R dysfunction in humans causes early-onset hyperphagia, impaired satiety and obesity. We have identified a novel c.216C>A (p.Asn72Lys) homozygous mutation in MC4R in a female child of Pakistani parents with no known consanguinity. The patient presented with early-onset severe obesity and hyperphagia. Developmental milestones were normal. On physical examination, height was 104.4 cm (+3 SD), weight 30 kg, and BMI 30 kg/m2 (+5.2 SD).  The mutation has not been described before but derives a high Grantham score of 94, indicating it is a likely cause for the phenotype of the subject. The mutation is in the predicted first cytoplasmic loop of MC4R. Only one other mutation has been described in this region of the receptor (p.H76R), which causes constitutive activation but decreased cell surface expression linking it with an obese phenotype. To confirm a causal link between the Asn72Lys mutation and the obese phenotype of the patient, we have undertaken a functional analysis of the receptor using in vitro methods. We have used site-directed mutagenesis to generate wild type and mutant MC4R expression constructs in the pcDNA3.1 plasmid. These were then transfected into HEK293 cells and assessed for responsiveness to α-MSH using Glo-Sensor technology to assess intra-cellular cAMP levels. We found that the maximum response (Rmax) of Asn72Lys-MC4R transfected cells was decreased to 20±1% of cells expressing wild type MC4R.  Moreover, the EC50 was increased from 15.4±1.2 nM to 35.0±1.3 nM compared to wild type MC4R transfectants. In conclusion, we describe a novel inactivating mutation in a new region of MC4R that significantly impairs its function and is associated with early-onset obesity and hyperphagia.

Nothing to Disclose: PJD, EB, MH, RV, APT, AJV, ELV

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