Park Jeenah, Sharma Neeraj, Cutting Garry R
McKusick-Nathans Institute of Genetic Medicine (J.P., N.S., G.R.C.), Johns Hopkins University, Baltimore, Maryland 21218; and Department of Pediatrics (G.R.C.), Johns Hopkins University School of Medicine, Baltimore, Maryland 21287-3914.
Mol Endocrinol. 2014 Sep;28(9):1547-57. doi: 10.1210/me.2014-1105. Epub 2014 Jul 22.
Melanocortin-3 receptor (MC3R) is a canonical MSH receptor that plays an essential role in energy homeostasis. Variants in MC3R have been implicated in obesity in humans and mice. However, interpretation of the functional consequences of these variants is challenging because the translational start site of MC3R is unclear. Using 5' rapid amplification of cDNA ends, we discovered a novel upstream exon that extends the length of the 5' untranslated region (UTR) in MC3R without changing the open-reading frame. The full-length 5' UTR directs utilization of an evolutionarily conserved second in-frame ATG as the primary translation start site. MC3R synthesized from the second ATG is localized to apical membranes of polarized Madin-Darby canine kidney cells, consistent with its function as a cell surface mediator of melanocortin signaling. Expression of MC3R causes relocalization of melanocortin receptor accessory protein 2, an accessory factor for melanocortin-2 receptor, to the apical membrane, coincident with the location of MC3R. In contrast, protein synthesized from MC3R cDNAs lacking the 5' UTR displayed diffuse cytosolic distribution and has no effect on the distribution of melanocortin receptor accessory protein 2. Our findings demonstrate that a previously unannotated 5' exon directs translation of MC3R protein that localizes to apical membranes of polarized cells. Together, our work provides insight on the structure of human MC3R and reveals a new pathway for regulation of energy metabolism.
黑皮质素-3受体（MC3R）是一种典型的促黑素受体，在能量稳态中起重要作用。MC3R基因变异与人类和小鼠的肥胖有关。然而，由于MC3R的翻译起始位点尚不清楚，对这些变异的功能后果进行解释具有挑战性。通过5' cDNA末端快速扩增技术，我们发现了一个新的上游外显子，它延长了MC3R中5'非翻译区（UTR）的长度，而不改变开放阅读框。全长5' UTR指导利用进化保守的第二个框内ATG作为主要翻译起始位点。从第二个ATG合成的MC3R定位于极化的Madin-Darby犬肾细胞的顶端膜，与其作为黑皮质素信号细胞表面介质的功能一致。MC3R的表达导致黑皮质素受体辅助蛋白2（黑皮质素-2受体的辅助因子）重新定位于顶端膜，与MC3R的位置一致。相比之下，缺乏5' UTR的MC3R cDNA合成的蛋白质表现出弥漫性的胞质分布，并且对黑皮质素受体辅助蛋白2的分布没有影响。我们的研究结果表明，一个以前未注释的5'外显子指导MC3R蛋白的翻译，该蛋白定位于极化细胞的顶端膜。总之，我们的工作为人类MC3R的结构提供了见解，并揭示了一条调节能量代谢的新途径。
