Heteromultimerization modulates P2X receptor functions through participating extracellular and C-terminal subdomains.
P2X purinergic receptors (P2XRs) differ among themselves with respect to their ligand preferences and channel kinetics during activation, desensitization, and recovery. However, the contributions of distinct receptor subdomains to the subtype-specific behavior have been incompletely characterized. Here we show that homomeric receptors having the extracellular domain of the P2X(3) subunit in the P2X(2a)-based backbone (P2X(2a)/X(3)ex) mimicked two intrinsic functions of P2X(3)R, sensitivity to alphabeta-methylene ATP and ecto-ATPase-dependent recovery from endogenous desensitization; these two functions were localized to the N- and C-terminal halves of the P2X(3) extracellular loop, respectively. The chimeric P2X(2a)R/X(3)ex receptors also desensitized with accelerated rates compared with native P2X(2a)R, and the introduction of P2X(2) C-terminal splicing into the chimeric subunit (P2X(2b)/X(3)ex) further increased the rate of desensitization. Physical and functional heteromerization of native P2X(2a) and P2X(2b) subunits was also demonstrated. In heteromeric receptors, the ectodomain of P2X(3) was a structural determinant for ligand selectivity and recovery from desensitization, and the C terminus of P2X(2) was an important factor for the desensitization rate. Furthermore, [gamma-(32)P]8-azido ATP, a photoreactive agonist, was effectively cross-linked to P2X(3) subunit in homomeric receptors but not in heteromeric P2X(2) + P2X(3)Rs. These results indicate that heteromeric receptors formed by distinct P2XR subunits develop new functions resulting from integrative effects of the participating extracellular and C-terminal subdomains.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001081:Apyrase / Q000378:metabolism / Q000494:pharmacology; D001386:Azides / Q000494:pharmacology; D019921:Biotinylation; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000737:chemistry; D007477:Ions; D007532:Isoleucine / Q000737:chemistry; D007700:Kinetics; D008024:Ligands; D051379:Mice; D009154:Mutation; D010649:Phenylalanine / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D018048:Receptors, Purinergic P2 / Q000737:chemistry* / Q000502:physiology*; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry; D014633:Valine / Q000737:chemistry; D014982:Xenopus laevis
D001386:Azides; D000939:Epitopes; D007477:Ions; D008024:Ligands; C546402:P2rx2 protein, mouse; C546409:P2rx3 protein, mouse; D018048:Receptors, Purinergic P2; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D007532:Isoleucine; D014443:Tyrosine; D010649:Phenylalanine; C017040:8-azidoadenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D001081:Apyrase; D014633:Valine; D002118:Calcium