Filipeanu Catalin M, Pullikuth Ashok K, Guidry Jessie J
Department of Pharmacology, College of Medicine, Howard University, Washington, DC (C.M.F.); Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, New Orleans, Louisiana (A.K.P., J.J.G.); Department of Cancer Biology, Wake Forest University School of Medicine, Winston Salem, North Carolina (A.K.P.); and Louisiana State University Health Sciences Center Proteomics Core Facility, New Orleans, Louisiana (J.J.G.)
Department of Pharmacology, College of Medicine, Howard University, Washington, DC (C.M.F.); Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, New Orleans, Louisiana (A.K.P., J.J.G.); Department of Cancer Biology, Wake Forest University School of Medicine, Winston Salem, North Carolina (A.K.P.); and Louisiana State University Health Sciences Center Proteomics Core Facility, New Orleans, Louisiana (J.J.G.).
Mol Pharmacol. 2015 May;87(5):792-802. doi: 10.1124/mol.114.096198. Epub 2015 Feb 13.
The human α2C-adrenergic receptor (α2C-AR) is localized intracellularly at physiologic temperature. Decreasing the environmental temperature strongly stimulates the receptor transport to the cell surface. In contrast, rat and mouse α2C-AR plasma membrane levels are less sensitive to decrease in temperature, whereas the opossum α2C-AR cell surface levels are not changed in these conditions. Structural analysis demonstrated that human α2C-AR has a high number of arginine residues in the third intracellular loop and in the C-terminus, organized as putative RXR motifs. Although these motifs do not affect the receptor subcellular localization at 37°C, deletion of the arginine clusters significantly enhanced receptor plasma membrane levels at reduced temperature. We found that this exaggerated transport of the human receptor is mediated by two functional arginine clusters, one in the third intracellular loop and one in the C-terminus. This effect is mediated by interactions with COPI vesicles, but not by 14-3-3 proteins. In rat α2C-AR, the arginine cluster from the third intracellular loop is shifted to the left due to three missing residues. Reinsertion of these residues in the rat α2C-AR restored the same temperature sensitivity as in the human receptor. Proteomic and coimmunoprecipitation experiments identified pontin as a molecule having stronger interactions with human α2C-AR compared with rat α2C-AR. Inhibition of pontin activity enhanced human receptor plasma membrane levels and signaling at 37°C. Our results demonstrate that human α2C-AR has a unique temperature-sensitive traffic pattern within the G protein-coupled receptor class due to interactions with different molecular chaperones, mediated in part by strict spatial localization of specific arginine residues.
人α2C - 肾上腺素能受体（α2C - AR）在生理温度下定位于细胞内。降低环境温度会强烈刺激该受体向细胞表面转运。相比之下，大鼠和小鼠α2C - AR的质膜水平对温度降低不太敏感，而负鼠α2C - AR的细胞表面水平在这些条件下没有变化。结构分析表明，人α2C - AR在第三个细胞内环和C末端有大量精氨酸残基，组织成假定的RXR基序。尽管这些基序在37°C时不影响受体的亚细胞定位，但精氨酸簇的缺失显著提高了低温下受体的质膜水平。我们发现，人受体这种过度的转运是由两个功能性精氨酸簇介导的，一个在第三个细胞内环，一个在C末端。这种效应是通过与COPI囊泡的相互作用介导的，而不是由14 - 3 - 3蛋白介导的。在大鼠α2C - AR中，由于三个缺失的残基，来自第三个细胞内环的精氨酸簇向左移动。将这些残基重新插入大鼠α2C - AR中恢复了与人受体相同的温度敏感性。蛋白质组学和免疫共沉淀实验确定pontin是一种与人α2C - AR相比与大鼠α2C - AR具有更强相互作用的分子。抑制pontin活性可提高人受体在37°C时的质膜水平和信号传导。我们的结果表明，由于与不同分子伴侣的相互作用，人α2C - AR在G蛋白偶联受体类别中具有独特的温度敏感运输模式，这部分是由特定精氨酸残基的严格空间定位介导的。
