Free fatty acids and protein kinase C activation induce GPR120 (free fatty acid receptor 4) phosphorylation.
GPR120, free fatty acid receptor 4, is a recently deorphanized G protein-coupled receptor that seems to play cardinal roles in the regulation of metabolism and in the pathophysiology of inflammatory and metabolic disorders. In the present work a GPR120-Venus fusion protein was expressed in HEK293 Flp-In T-REx cells and its function (increase in intracellular calcium) and phosphorylation were studied. It was observed that the fusion protein migrated in sodium dodecyl sulfate-polyacrylamide gels as a band with a mass of ≈70-75kDa, although other bands of higher apparent weight (>130kDa) were also detected. Cell stimulation with docosahexaenoic acid or α-linolenic acid induced concentration-dependent increases in intracellular calcium and GPR120 phosphorylation. Activation of protein kinase C with phorbol esters also induced a marked receptor phosphorylation but did not alter the ability of 1µM docosahexaenoic acid to increase the intracellular calcium concentration. Phorbol ester-induced GPR120 phosphorylation, but not that induced with docosahexaenoic acid, was blocked by protein kinase C inhibitors (bis-indolyl-maleimide I and Gö 6976) suggesting that conventional kinase isoforms mediate this action. The absence of effect of protein kinase C inhibitors on agonist-induced GPR120 phosphorylation indicates that this kinase does not play a major role in agonist-induced receptor phosphorylation. Docosahexaenoic acid action was associated with marked GPR120 internalization whereas that induced with phorbol esters was smaller at early times.
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Ap. Postal 70-248, México D. F. 04510, Mexico.
Omar B
OB
Sánchez-Reyes
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Ap. Postal 70-248, México D. F. 04510, Mexico.
M Teresa
MT
Romero-Ávila
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Ap. Postal 70-248, México D. F. 04510, Mexico.
Jean A
JA
Castillo-Badillo
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Yoshinori
Y
Takei
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Akira
A
Hirasawa
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Gozoh
G
Tsujimoto
Unidad de Biomedicina, Facultad de Estudios Superiores Iztacala, Universidad Nacional Autónoma de México, Tlalnepantla, Mexico.
Rafael
R
Villalobos-Molina
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Ap. Postal 70-248, México D. F. 04510, Mexico. Electronic address: agarcia@ifc.unam.mx.
J Adolfo
JA
García-Sáinz
D004281:Docosahexaenoic Acids / Q000494:pharmacology*; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D017962:alpha-Linolenic Acid / Q000494:pharmacology*
C519853:FFAR4 protein, human; D005230:Fatty Acids, Nonesterified; D010703:Phorbol Esters; D043562:Receptors, G-Protein-Coupled; D011993:Recombinant Fusion Proteins; D017962:alpha-Linolenic Acid; D004281:Docosahexaenoic Acids; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate
Free fatty acid receptor 4; GPR120; Protein kinase C; Receptor phosphorylation