Molecular Determinants of the Human α_2C-Adrenergic Receptor Temperature-Sensitive Intracellular Traffic

Abstract

The human a 2C -adrenergic receptor (a 2C -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 a 2C -AR plasma membrane levels are less sensitive to decrease in temperature, whereas the opossum a 2C -AR cell surface levels are not changed in these conditions. Structural analysis demonstrated that human a 2C -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 a 2C -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 a 2C -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 a 2C -AR compared with rat a 2C -AR. Inhibition of pontin activity enhanced human receptor plasma membrane levels and signaling at 37°C. Our results demonstrate that human a 2C -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.

The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

“…This subtle adaptation to the temperature of the upper or lower airways is also evident for influenza virus hemagglutinin [13], suggesting that it might be a commonality for human respiratory viruses. As to the biochemical basis, preference for 33˚C for virus production indicates that the S protein is relatively unstable, since a cooler temperature should reduce conformational flexibility during S glycoprotein synthesis or transport [69], and avoid formation of unstable spike conformers. Although the spike's temperature dependency may seem quite subtle, its relevance becomes clear from our comparison of the two SARS-2-S variants.…”

The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

“…This airway temperature preference is a property of the coronavirus spike glycoprotein, but also the influenza virus hemagglutinin (13), suggesting that it might be a commonality for human respiratory viruses. As to the biochemical basis, a cooler temperature likely ensures lower conformational flexibility during S glycoprotein synthesis or transport (60), and avoidance of unstable spike conformers. For SARS-2-S, the 33°C preference proved more pronounced for the S D614 than the S G614 variant.…”

A single mutation in helix 8 enhances the angiotensin II type 1a receptor transport and signaling

“…The lack of response of the AT1aR mutant K308E to Rab6 inhibition suggests that Lys308 may be involved in the regulation of AT1aR retrograde movement. It is also interesting to note that, in some GPCRs such as α 2C -adrenergic and GABA B receptors, positively charged residues may function as ER retention motifs to block the receptor export from the ER [56, 57]. To the best of our knowledge, these data provide the first evidence implicating a role of retrograde transport in regulating GPCR forward trafficking.…”