Stimulation of increases in intracellular calcium and prostaglandin E2 generation in Chinese hamster ovary cells expressing receptor-Galpha16 fusion proteins

• Published in: Journal of biochemistry (Tokyo) Vol. 135; no. 5; pp. 605 - 613
• Main Authors: Suga, Hinako, Takeda, Shigeki, Haga, Tatsuya, Okamura, Michiko, Takao, Kyoichi, Tatemoto, Kazuhiko
• Format: Journal Article
• Language: English
• Published: England 01.05.2004
• Subjects: Animals; Calcium - metabolism; Cell Line; CHO Cells; Cricetinae; CX3C Chemokine Receptor 1; Dinoprostone - metabolism; DNA, Complementary - metabolism; Dose-Response Relationship, Drug; GTP-Binding Protein alpha Subunits, Gq-G11; Heterotrimeric GTP-Binding Proteins - chemistry; Heterotrimeric GTP-Binding Proteins - metabolism; Humans; Insecta; Ligands; Membrane Proteins - metabolism; Plasmids - metabolism; Prostaglandins - metabolism; Protein Binding; Receptor, Muscarinic M2 - metabolism; Receptors, Chemokine - metabolism; Receptors, G-Protein-Coupled - metabolism; Recombinant Fusion Proteins - metabolism; Reverse Transcriptase Polymerase Chain Reaction; Time Factors; Transfection
• ISSN: 0021-924X

We examined whether fusion proteins of G protein-coupled receptors with the alpha subunit of G(16) (Galpha(16)) could activate downstream signals. We expressed fusion proteins of G(i)-coupled receptors, i.e. CX(3)C chemokine receptor 1 (CX(3)CR1) and M(2) receptor, in Chinese hamster ovary cells. An agonist for CX(3)CR1 induced greater increases in intracellular Ca(2+) and prostaglandin E(2) generation in cells expressing CX(3)CR1-Galpha(16) fusion protein than in cells expressing CX(3)CR1 alone or both CX(3)CR1 and Galpha(16) separately. Similarly, agonist-induced prostaglandin E(2) generation was greater in cells expressing M(2)-Galpha(16) fusion protein than ones expressing M(2) alone or both M(2) and Galpha(16) separately. In cells expressing fusion proteins with Galpha(16) of G(q)-coupled receptors, i.e. urotensin II receptor and M(1) receptor, the relevant agonists induced similar increases in intracellular Ca(2+) and prostaglandin E(2) generation as in ones expressing the receptor alone. In cells expressing urotensin II receptor-Galpha(16) fusion protein, prostaglandin E(2) generation exhibited a lower EC(50) value than the intracellular Ca(2+) increase. These results indicate that agonist-stimulated receptor-Galpha(16) fusion proteins are coupled to downstream signaling pathways, and suggest that receptor-Galpha(16) fusion proteins may be useful for screening for ligands of orphan G protein-coupled receptors and G(i)-coupled receptors.