| First Author | Van Koppen CJ | Year | 1994 |
| Journal | Eur J Biochem | Volume | 222 |
| Issue | 2 | Pages | 525-31 |
| PubMed ID | 8020490 | Mgi Jnum | J:19537 |
| Mgi Id | MGI:67694 | Doi | 10.1111/j.1432-1033.1994.tb18894.x |
| Citation | Van Koppen CJ, et al. (1994) Deletion analysis of the m4 muscarinic acetylcholine receptor. Molecular determinants for activation of but not coupling to the Gi guanine-nucleotide-binding regulatory protein regulate receptor internalization. Eur J Biochem 222(2):525-31 |
| abstractText | In order to investigate whether coupling to and/or activation of guanine-nucleotide-binding proteins (G proteins) is involved in agonist-induced internalization of m4 muscarinic acetylcholine receptors (mAChRs), a deletion mutant [des-(264-394)mAChR] was constructed that lacks a substantial portion of the putative third intracellular loop. The wild-type receptor and des-(264-394)mAChR stably expressed in Chinese hamster ovary cells in essentially comparable amounts, exhibited identical antagonist-binding affinities. Consistent with the reported importance of the third cytoplasmic loop for Gi protein activation, the des-(264-394)mAChR showed a drastically reduced potential to mediate agonist-induced inhibition of adenylyl cyclase. In contrast, the ability of the mutant receptor to couple to Gi proteins was not impaired, as demonstrated by a similar guanine-nucleotide-sensitive and pertussis-toxin-sensitive high-affinity agonist-receptor binding for both mAChRs. In contrast, des-(264-394)mAChR was hardly able to stimulate the GTPase activity of G proteins, suggesting impaired activation of Gi proteins rather than ineffective coupling to Gi proteins. Internalization of wild-type receptor and des-(264-394)mAChR was observed with similar agonist concentrations and showed similar maximal values. However, des-(264-394)mAChR displayed a significantly reduced rate of receptor internalization. A similar attenuation of wild-type mAChR internalization was obtained upon pertussis toxin treatment. In conclusion, our data provide evidence that the molecular determinants of the m4 mAChR involved in Gi-protein coupling and activation are not identical and that activation of, but not coupling to, Gi proteins regulates m4 mAChR internalization. |
