Phosphorylation of Galpha11 protein contributes to agonist-induced desensitization of 5-HT2A receptor signaling.
Agonist treatment causes desensitization of many G protein-coupled receptor systems. Recent advances have delineated changes in receptors in the desensitization response; however, the role of G proteins remains unclear. We investigated the role of phosphorylation of Galpha q/11 proteins in agonist-induced desensitization of serotonin 2A (5-HT2A) receptors. In an embryonic rat cortical cell line (A1A1v), 24-h treatment with 100 nM (-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl (DOI), a 5-HT(2A/2C) receptor agonist, decreased DOI-stimulated inositol phosphate accumulation and increased the phosphorylation of Galpha q/11 proteins, as demonstrated by immunoprecipitation of Galpha q/11 and both incorporation of 32P phosphate and labeling with a S/T/Y phosphorylation-dependent antibody. Treatment with DOI for 30 min induced desensitization but did not increase phosphorylation of Galpha q/11 proteins, suggesting that different mechanisms are involved in desensitization after short- and long-term treatments. Mutation of S154A in a protein kinase C (PKC) and calcium/calmodulin dependent kinase (CaMK) consensus site in Galpha11 significantly reduced DOI-stimulated phosphorylation of Galpha11 and DOI-induced desensitization of 5-HT2A receptor signaling. Inhibition of PKC and CaMK attenuated phosphorylation of Galpha q/11 proteins and DOI-induced desensitization of 5-HT2A receptors. Expression of Galpha11 S154D, a phosphorylation mimic, reduced DOI-stimulated inositol phosphate accumulation. DOI treatment for 24 h also produced heterologous desensitization, as indicated by decreased bradykinin-stimulated inositol phosphate accumulation. These data suggest that phosphorylation of Galpha11 protein by PKC and CaMK contributes to agonist-induced homologous desensitization of 5-HT2A receptor signaling as well as heterologous desensitization. The phosphorylation of Galpha protein represents a novel mechanism involved in regulation of receptor signaling and agonist-induced desensitization of G protein-coupled receptors.
Shi
Ju
J
Department of Pharmacology Loyola University Chicago, Stritch School of Medicine, 2160 South First Avenue, Maywood, IL 60153, USA.
Zemaitaitis
Bozena
B
Muma
Nancy A
NA
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002460:Cell Line; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009474:Neurons; D010766:Phosphorylation; D020641:Polymorphism, Single Nucleotide; D044402:Receptor, Serotonin, 5-HT2A / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine; D015398:Signal Transduction; D013912:Threonine; D014162:Transfection
D044402:Receptor, Serotonin, 5-HT2A; D011994:Recombinant Proteins; D013912:Threonine; D012694:Serine; D043802:GTP-Binding Protein alpha Subunits, Gq-G11