Alternative splicing of the dopamine D2 receptor directs specificity of coupling to G-proteins.
Two isoforms of the dopamine D2 receptor have been characterized, D2L (long) and D2S (short), generated by alternative splicing from the same gene. They differ by an in-frame insert of 29 amino acids specific to D2L within the putative third intracytoplasmic loop of the receptor. We have previously demonstrated (Montmayeur, J.-P., Guiramand, J., and Borelli, E. (1993) Mol. Endocrinol. 7, 161-170) that D2S and D2L, although presenting very similar pharmacological profiles, couple differently to the alpha-subunit of guanine nucleotide-binding regulatory proteins (G-proteins). In particular, D2L, but not D2S, requires the presence of the alpha-subunit of the inhibitory G-protein (G alpha i2) to elicit greater inhibition of adenylyl cyclase activity. The insert present in D2L must therefore confer the specificity of interaction with G alpha i2. Thus, we introduced substitution mutations within the D2L insert. These mutant receptors were expressed in JEG3 cells, a G alpha i2-deficient cell line, scoring for those presenting an increased inhibition of adenylyl cyclase by dopamine. Our analysis identified two mutants, S259/262A and D249V, with these properties. These results clearly show that the insert present in D2L plays a critical role in the selectivity for the G-proteins interacting with the receptor.
Guiramand
J
J
Institut de Génétique et de Biologie Moléculaire et Cellulaire, U.184 INSERM/CNRS/Université Louis Pasteur, Illkirch, France.
Montmayeur
J P
JP
Ceraline
J
J
Bhatia
M
M
Borrelli
E
E
D000067956:Adenylyl Cyclase Inhibitors; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001058:Apomorphine / Q000031:analogs & derivatives / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000378:metabolism; D004298:Dopamine / Q000494:pharmacology; D018491:Dopamine Agonists / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007545:Isoproterenol / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D018343:Receptors, Adrenergic, beta-2 / Q000096:biosynthesis / Q000378:metabolism; D017448:Receptors, Dopamine D2 / Q000096:biosynthesis* / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D013134:Spiperone / Q000378:metabolism; D014162:Transfection
D000067956:Adenylyl Cyclase Inhibitors; D018491:Dopamine Agonists; D018343:Receptors, Adrenergic, beta-2; D017448:Receptors, Dopamine D2; D011994:Recombinant Proteins; D013134:Spiperone; C009074:N-n-propylnorapomorphine; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D007545:Isoproterenol; D001058:Apomorphine; D004298:Dopamine