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P2X Receptor Chimeras Highlight Roles of the Amino Terminus to Partial Agonist Efficacy, the Carboxyl Terminus to Recovery from Desensitization, and Independent Regulation of Channel Transitions

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journal contribution
posted on 2013-10-17, 15:39 authored by Rebecca C. Allsopp, Louise K. Farmer, Alistair G. Fryatt, Richard J. Evans
P2X receptor subtypes can be distinguished by their sensitivity to ATP analogues and selective antagonists. We have used chimeras between human P2X1 and P2X2 receptors to address the contribution of the extracellular ligand binding loop, transmembrane segments (TM1 and TM2), and intracellular amino and carboxyl termini to the action of partial agonists (higher potency and efficacy of BzATP andAp5Aat P2X1 receptors) and antagonists. Sensitivity to the antagonists NF449, suramin, and PPADS was conferred by the nature of the extracellular loop (e.g. nanomolar for NF449 at P2X1 and P2X2-1EXT and micromolar at P2X2 and P2X1-2EXT). In contrast, the effectiveness of partial agonists was similar to P2X1 levels for both of the loop transfers, suggesting that interactions with the rest of the receptor played an important role. Swapping TM2 had reciprocal effects on partial agonist efficacy. However, TM1 swaps increased partial agonist efficacy at both chimeras, and this was similar for swaps of both TM1 and 2. Changing the amino terminus had no effect on agonist potency but increased partial agonist efficacy at P2X2-1N and decreased it at P2X1-2N chimeras, demonstrating that potency and efficacy can be independently regulated. Chimeras and point mutations also identified residues in the carboxyl terminus that regulated recovery from channel desensitization. These results show that interactions among the intracellular, transmembrane, and extracellular portions of the receptor regulate channel properties and suggest that transitions to channel opening, the behavior of the open channel, and recovery from the desensitized state can be controlled independently.

Funding

This work was supported by the Wellcome Trust and the British Heart Foundation.

History

Citation

Journal of Biological Chemistry, 2013, 288 (29), pp. 21412–21421

Author affiliation

/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Biological Sciences/Department of Cell Physiology and Pharmacology

Version

  • VoR (Version of Record)

Published in

Journal of Biological Chemistry

Publisher

The American Society for Biochemistry and Molecular Biology, Inc.

issn

0021-9258

eissn

1083-351X

Copyright date

2013

Available date

2013-10-17

Publisher version

http://www.jbc.org/content/288/29/21412.full

Language

en