Differential regulation of muscarinic M1 and M3 receptors by a putative phosphorylation domain.
A motif consisting of several serine residues flanked N-terminally by acidic residues occurs in the third intracellular loop of both muscarinic M1 and M3 receptors (287SerLeuThrSerSer291 and 349SerAlaSerSer352, respectively). We examined the role of these domains in modulating agonist-induced desensitization and receptor trafficking, and for the muscarinic M3 receptor, we assessed the contribution of phosphorylation to receptor regulation. Mutation of the above residues did not affect desensitization of phosphoinositide hydrolysis signaling for either the muscarinic M1 or M3 receptor and did not alter the agonist-induced phosphorylation state of the muscarinic M3 receptor. Mutation of this domain (349SerAlaSerSer352/349AlaAlaAlaAla352) in the muscarinic M3 receptor completely abrogated receptor internalization and subsequently, down-regulation. Mutation of the analogous domain (287SerLeuThrSerSer291/287AlaLeuAlaAlaAla291) in the muscarinic M1 receptor had no obvious effect on internalization, but led to a more rapid down-regulation. Thus, these serine-rich regions are not required for receptor desensitization, but are differentially involved in receptor trafficking for the muscarinic M1 and M3 receptors.
Shockley
M S
MS
Department of Biopharmaceutical Sciences, University of California, San Francisco 94143, USA.
Tolbert
L M
LM
Tobin
A B
AB
Nahorski
S R
SR
Sadée
W
W
Lameh
J
J
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D016466:CHO Cells; D002217:Carbachol / Q000494:pharmacology; D006224:Cricetinae; D015536:Down-Regulation; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D018613:Microscopy, Confocal; D018721:Muscarinic Agonists / Q000494:pharmacology; D009154:Mutation; D010766:Phosphorylation / Q000187:drug effects; D011869:Radioligand Assay; D043643:Receptor, Muscarinic M1; D043587:Receptor, Muscarinic M3; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D018721:Muscarinic Agonists; D043643:Receptor, Muscarinic M1; D043587:Receptor, Muscarinic M3; D011956:Receptors, Cell Surface; D011976:Receptors, Muscarinic; D015544:Inositol 1,4,5-Trisphosphate; D002217:Carbachol