Functional characterization of mutations in melanocortin-4 receptor associated with human obesity
Functional characterization of mutations in melanocortin-4 receptor associated with human obesity
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DOI：
10.1074/jbc.274.50.35816
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发表时间：
1999-12-10
影响因子：
4.8
通讯作者：
MacKenzie, RG
中科院分区：
文献类型：
--
作者：
Ho, GY;MacKenzie, RG
黑皮质素-4受体（MC4R）是一种G蛋白偶联受体，参与体重调节。人类的遗传研究已经确定了MC4R的两个移码突变与显性遗传形式的肥胖有关。我们已经在293 T细胞中产生并表达了相应的MC4R突变体，并且发现用截短突变体转染的细胞未能表现出激动剂结合或响应性，尽管保留了可能足以用于结合和信号传导的结构基序。免疫荧光研究表明，突变蛋白的表达和定位在细胞内室，但从质膜缺席，这表明这些突变破坏了适当的细胞转运MC4R。进一步的研究鉴定了MC4R细胞质尾区中细胞表面靶向所必需的序列。我们进一步研究了突变体对野生型受体功能的可能的显性负活性。共转染研究表明，突变体既不影响信号传导，也不影响野生型MC4R的细胞表面表达。我们还表征了MC4R的三种人类序列变体，但这些变体对肽配体表现出相同的亲和力和相同的激动剂反应性。因此，与肥胖相关的MC4R截短突变体不同，MC4R的多态性不太可能是人类肥胖的贡献者。
Melanocortin-4 receptor (MC4R) is a G protein-coupled receptor implicated in the regulation of body weight. Genetic studies in humans have identified two frameshift mutations of MC4R associated with a dominantly inherited form of obesity. We have generated and expressed the corresponding MC4R mutants in 293T cells and found that cells transfected with the truncation mutants failed to exhibit agonist binding or responsiveness despite retention of structural motifs potentially sufficient for binding and signaling. Immunofluorescence studies showed that the mutant proteins were expressed and localized in the intracellular compartment but absent from the plasma membrane, suggesting that these mutations disrupted the proper cellular transport of MC4R. Further studies identified a sequence in the cytoplasmic tail of MC4R necessary for the cell surface targeting. We further investigated a possible dominant-negative activity of the mutants on wildtype receptor function. Co-transfection studies showed that the mutants affected neither signaling nor cell surface expression of wild-type MC4R. We also characterized three human sequence variants of MC4R, but these exhibited identical affinities for peptide ligands and identical agonist responsiveness. Thus, unlike the obesity-associated MC4R truncation mutants, the polymorphisms of MC4R are unlikely to be contributors to human obesity.