Intracellular retention of human melanocortin-4 receptor: a molecular mechanism underlying early-onset obesity in F261S pedigree of Chinese.

Objective: To investigate how F261S mutation identified from Chinese obese patients affects the function of melanocortin 4 receptor (MC4R) and to analyze the obesity-related phenotypes in subjects carrying the F261S mutation.

Methods: F261S mutant of MC4R was generated by site-directed mutagenesis. Plasmids encoding wild-type or F261S mutant of MC4R were transfected into HEK293 and COS-7 cells to examine their functional characteristics. Signaling properties of F261S MC4R were assessed by measuring intracellular cAMP levels in response to alpha-MSH stimulation. Cell surface expression of F261S MC4R was compared with that of wild-type MC4R. Clinical examinations were performed in subjects carrying F261S mutation and in non-mutated controls.

Results: The alpha-MSH-stimulated reporter gene activity was significantly reduced in cells expressing F261S MC4R, with a maximal response equal to 57% of wild-type MC4R. The F261S mutation also led to a significant change in the Es50 value compared with the wild-type receptor (P<0.01). Immunofluorescent assay revealed a marked reduction in plasma membrane localization of the MC4R in cells expressing the F261S mutant receptor. The resting metabolic rate and fat composition of the mutant carriers were not significantly different from those of the non-mutated obese controls.

Conclusions: The decreased response to alpha-MSH due to the intracellular retention of MC4R may cause early-onset obesity in the F261S pedigree of Chinese.