Beta-adrenergic receptor stimulation promotes G alpha s internalization through lipid rafts: a study in living cells - PubMed

Affiliations

• PMID: 15703379
• DOI: 10.1124/mol.104.008342

Comparative Study

Beta-adrenergic receptor stimulation promotes G alpha s internalization through lipid rafts: a study in living cells

John A Allen et al. Mol Pharmacol. 2005 May.

Abstract

Upon binding hormones or drugs, many G protein-coupled receptors are internalized, leading to receptor recycling, receptor desensitization, and down-regulation. Much less understood is whether heterotrimeric G proteins also undergo agonist-induced endocytosis. To investigate the intracellular trafficking of G alpha s, we developed a functional G alpha s-green fluorescent protein (GFP) fusion protein that can be visualized in living cells during signal transduction. C6 and MCF-7 cells expressing G alpha s-GFP were treated with 10 microM isoproterenol, and trafficking was assessed with fluorescence microscopy. Upon isoproterenol stimulation, G alpha s-GFP was removed from the plasma membrane and internalized into vesicles. Vesicles containing G alpha s-GFP did not colocalize with markers for early endosomes or late endosomes/lysosomes, revealing that G alpha s does not traffic through common endocytic pathways. Furthermore, G alpha s-GFP did not colocalize with internalized beta2-adrenergic receptors, suggesting that G alpha s and receptors are removed from the plasma membrane by distinct endocytic pathways. Nonetheless, activated G alpha s-GFP did colocalize in vesicles labeled with fluorescent cholera toxin B, a lipid raft marker. Agonist significantly increased G alpha s protein in Triton X-100 -insoluble membrane fractions, suggesting that G alpha s moves into lipid rafts/caveolae after activation. Disruption of rafts/caveolae by treatment with cyclodextrin prevented agonist-induced internalization of G alpha s-GFP, as did overexpression of a dominant-negative dynamin. Taken together, these results suggest that receptor-activated G alpha s moves into lipid rafts and is internalized from these membrane microdomains. It is suggested that agonist-induced internalization of G alpha s plays a specific role in G protein-coupled receptor-mediated signaling and could enable G alpha s to traffic into the cellular interior to regulate effectors at multiple cellular sites.

Similar articles

• Mastoparan inhibits beta-adrenoceptor-G(s) signaling by changing the localization of Galpha(s) in lipid rafts.

  Sugama J, Yu JZ, Rasenick MM, Nakahata N. Sugama J, et al. Cell Signal. 2007 Nov;19(11):2247-54. doi: 10.1016/j.cellsig.2007.06.024. Epub 2007 Jul 4. Cell Signal. 2007. PMID: 17692506

• Differences in endosomal targeting of human (beta)1- and (beta)2-adrenergic receptors following clathrin-mediated endocytosis.

  Liang W, Curran PK, Hoang Q, Moreland RT, Fishman PH. Liang W, et al. J Cell Sci. 2004 Feb 15;117(Pt 5):723-34. doi: 10.1242/jcs.00878. Epub 2004 Jan 20. J Cell Sci. 2004. PMID: 14734649

• Fas signaling induces raft coalescence that is blocked by cholesterol depletion in human RPE cells undergoing apoptosis.

  Lincoln JE, Boling M, Parikh AN, Yeh Y, Gilchrist DG, Morse LS. Lincoln JE, et al. Invest Ophthalmol Vis Sci. 2006 May;47(5):2172-8. doi: 10.1167/iovs.05-1167. Invest Ophthalmol Vis Sci. 2006. PMID: 16639029

• Caveosomes and endocytosis of lipid rafts.

  Nichols B. Nichols B. J Cell Sci. 2003 Dec 1;116(Pt 23):4707-14. doi: 10.1242/jcs.00840. J Cell Sci. 2003. PMID: 14600257 Review.

• Stoichiometry and compartmentation in G protein-coupled receptor signaling: implications for therapeutic interventions involving G(s).

  Ostrom RS, Post SR, Insel PA. Ostrom RS, et al. J Pharmacol Exp Ther. 2000 Aug;294(2):407-12. J Pharmacol Exp Ther. 2000. PMID: 10900212 Review.

Cited by

• Subcellular localization of mu-opioid receptor G(s) signaling.

  Chakrabarti S, Chang A, Gintzler AR. Chakrabarti S, et al. J Pharmacol Exp Ther. 2010 Apr;333(1):193-200. doi: 10.1124/jpet.109.165142. Epub 2010 Jan 22. J Pharmacol Exp Ther. 2010. PMID: 20097777 Free PMC article.

• Disruption of lipid-raft localized Gα_s/tubulin complexes by antidepressants: a unique feature of HDAC6 inhibitors, SSRI and tricyclic compounds.

  Singh H, Wray N, Schappi JM, Rasenick MM. Singh H, et al. Neuropsychopharmacology. 2018 Jun;43(7):1481-1491. doi: 10.1038/s41386-018-0016-x. Epub 2018 Feb 5. Neuropsychopharmacology. 2018. PMID: 29463911 Free PMC article. Retracted.

• Differential expression of Gs in a murine model of homocysteinemic heart failure.

  Vacek TP, Sen U, Tyagi N, Vacek JC, Kumar M, Hughes WM, Passmore JC, Tyagi SC. Vacek TP, et al. Vasc Health Risk Manag. 2009;5(1):79-84. Epub 2009 Apr 8. Vasc Health Risk Manag. 2009. PMID: 19436674 Free PMC article.

• Submembraneous microtubule cytoskeleton: regulation of microtubule assembly by heterotrimeric Gproteins.

  Roychowdhury S, Rasenick MM. Roychowdhury S, et al. FEBS J. 2008 Oct;275(19):4654-63. doi: 10.1111/j.1742-4658.2008.06614.x. Epub 2008 Aug 27. FEBS J. 2008. PMID: 18754776 Free PMC article. Review.

• Gαs regulates the post-endocytic sorting of G protein-coupled receptors.

  Rosciglione S, Thériault C, Boily MO, Paquette M, Lavoie C. Rosciglione S, et al. Nat Commun. 2014 Aug 4;5:4556. doi: 10.1038/ncomms5556. Nat Commun. 2014. PMID: 25089012 Free PMC article.

Publication types

MeSH terms

Substances