ABSTRACT
P2X receptors (P2XRs) are trimeric ligand-gated ion channels that open a cation-selective pore in response to ATP binding to their large extracellular domain (ECD). The seven known P2XR subtypes typically assemble as homo- or heterotrimeric complexes and they contribute to numerous physiological functions, including nociception, inflammation and hearing. Both the overall structure of P2XRs and the details of how ATP is coordinated at the subunit interface are well established. By contrast, little is known about how inter-subunit interactions in the ECD contribute to channel function. Here we investigate both single and double mutants at the subunit interface of rP2X2Rs using electrophysiological and biochemical approaches. Our data demonstrate that the vast majority of mutations that disrupt putative inter-subunit interactions result in channels with higher apparent ATP affinity and that double mutants at the subunit interface show significant energetic coupling, especially if the mutations are located in close proximity. Overall, we show that inter-subunit interactions, as well as possibly interactions in other parts of the receptor, stabilize WT rP2X2Rs in the closed state. This suggests that, unlike other ligand-gated ion channels, P2X2 receptors have not evolved for an intrinsically low threshold for activation, possibly to allow for additional modulation or as a cellular protection mechanism against overstimulation.
Competing Interest Statement
The authors have declared no competing interest.
