guanosine-triphosphate and 9-hydroxy-10-12-octadecadienoic-acid

G2A is a G protein-coupled receptor that can be induced by various stressors. G2A is reported to have proton-sensing activity that mediates intracellular inositol phosphate (IP) accumulation with decreasing pH. We previously showed that G2A is also activated by some oxidized free fatty acids such as 9-hydroxyoctadecadienoic acid (9-HODE). In this study, we identified a novel alternative splice variant of G2A (G2A-b) that has a partially different N terminus compared with the G2A originally reported (G2A-a). The two splice variants of G2A show similar tissue distributions, but G2A-b is expressed more abundantly. There was no difference between the two variants in 9-HODE-induced cellular responses, such as intracellular calcium mobilization and GDP/GTP exchange of Galpha protein, and in proton-sensitive IP accumulation. However, G2A-b showed a higher basal activity in terms of IP accumulation. Mutagenesis study revealed that the difference in the basal activity is attributable to the K7 residue that exists only in G2A-a. We further demonstrated that an R42A mutation largely impaired both the basal and proton-sensing activities, but did not affect the 9-HODE-induced intracellular calcium increase. Taken together, we found an additional novel G2A variant (G2A-b) that is the major transcript with functional response to ligand stimulation as well as G2A-a, and succeeded in discriminating proton-sensing and oxidized fatty acid-sensing activities of G2A.

Topics: Alternative Splicing; Amino Acid Sequence; Animals; Calcium; Cell Cycle Proteins; Chlorocebus aethiops; CHO Cells; COS Cells; Cricetinae; Cricetulus; Cyclic AMP Response Element-Binding Protein A; Guanosine Triphosphate; HL-60 Cells; Humans; Inositol Phosphates; Leukocytes; Linoleic Acids, Conjugated; Molecular Sequence Data; Protein Isoforms; Receptors, G-Protein-Coupled; RNA, Messenger; Serum Response Element; Transfection