Heteromultimerization modulates P2X receptor functions through participating extracellular and C-terminal subdomains

Abstract

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 P2X3 subunit in the P2X2a-based backbone (P2X2a/X3ex) mimicked two intrinsic functions of P2X3R, sensitivity to αβ-methylene ATP and ecto-ATPase-dependent recovery from endogenous desensitization; these two functions were localized to the N- and C-terminal halves of the P2X3 extracellular loop, respectively. The chimeric P2X2aR/X3ex receptors also desensitized with accelerated rates compared with native P2X2aR, and the introduction of P2X2 C-terminal splicing into the chimeric subunit (P2X2b/X3ex) further increased the rate of desensitization. Physical and functional heteromerization of native P2X2a and P2X2b subunits was also demonstrated. In heteromeric receptors, the ectodomain of P2X3 was a structural determinant for ligand selectivity and recovery from desensitization, and the C terminus of P2X2 was an important factor for the desensitization rate. Furthermore, [γ-32P]8-azido ATP, a photoreactive agonist, was effectively cross-linked to P2X3 subunit in homomeric receptors but not in heteromeric P2X2 + P2X3Rs. 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.