Heteromeric association creates a P2Y-like adenosine receptor.
Adenosine and its endogenous precursor ATP are main components of the purinergic system that modulates cellular and tissue functions via specific adenosine and ATP receptors (P1 and P2 receptors), respectively. Although adenosine inhibits excitability and ATP functions as an excitatory transmitter in the central nervous system, little is known about the ability of P1 and P2 receptors to form new functional structures such as a heteromer to control the complex purinergic cascade. Here we have shown that G(i/o) protein-coupled A1 adenosine receptor (A1R) and Gq protein-coupled P2Y1 receptor (P2Y1R) coimmunoprecipitate in cotransfected HEK293T cells, suggesting the oligomeric association between distinct G protein-coupled P1 and P2 receptors. A1R and P2Y2 receptor, but not A1R and dopamine D2 receptor, also were found to coimmunoprecipitate in cotransfected cells. A1R agonist and antagonist binding to cell membranes were reduced by coexpression of A1R and P2Y1R, whereas a potent P2Y1R agonist adenosine 5'-O-(2-thiotriphosphate) (ADPbetaS) revealed a significant potency to A1R binding only in the cotransfected cell membranes. Moreover, the A1R/P2Y1R coexpressed cells showed an ADPbetaS-dependent reduction of forskolin-evoked cAMP accumulation that was sensitive to pertussis toxin and A1R antagonist, indicating that ADPbetaS binds A1R and inhibits adenylyl cyclase activity via G(i/o) proteins. Also, a high degree of A1R and P2Y1R colocalization was demonstrated in cotransfected cells by double immunofluorescence experiments with confocal laser microscopy. These results suggest that oligomeric association of A1R with P2Y1R generates A1R with P2Y1R-like agonistic pharmacology and provides a molecular mechanism for an increased diversity of purine signaling.
Yoshioka
K
K
Department of Molecular and Cellular Neurobiology, Tokyo Metropolitan Institute for Neuroscience, Fuchu, Tokyo 183-8526, Japan.
Saitoh
O
O
Nakata
H
H
D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D000242:Cyclic AMP / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000737:chemistry / Q000302:isolation & purification / Q000502:physiology; D020962:Heterotrimeric GTP-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000502:physiology; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D058906:Purinergic P1 Receptor Agonists; D011869:Radioligand Assay; D051381:Rats; D017448:Receptors, Dopamine D2 / Q000737:chemistry / Q000302:isolation & purification / Q000502:physiology; D018047:Receptors, Purinergic P1 / Q000302:isolation & purification / Q000502:physiology*; D018048:Receptors, Purinergic P2 / Q000737:chemistry / Q000302:isolation & purification / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014162:Transfection; D014316:Tritium; D014970:Xanthines / Q000493:pharmacokinetics
D046911:Macromolecular Substances; D058906:Purinergic P1 Receptor Agonists; D017448:Receptors, Dopamine D2; D018047:Receptors, Purinergic P1; D018048:Receptors, Purinergic P2; D011994:Recombinant Proteins; D014970:Xanthines; D014316:Tritium; C048599:N(6)-cyclopentyladenosine; D015544:Inositol 1,4,5-Trisphosphate; C051360:1,3-dipropyl-8-cyclopentylxanthine; D000242:Cyclic AMP; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D020962:Heterotrimeric GTP-Binding Proteins; D000262:Adenylyl Cyclases; D000241:Adenosine