Neuroscience 2004 Abstract
| Presentation Number: | 958.2 |
|---|---|
| Abstract Title: | Cytosolic and transmembrane domain gating motions associated with P2x2 channel permeability dynamics. |
| Authors: |
Khakh, B. S.*1
; Fisher, J.1
; Egan, T. M.
1MRC Lab. of Mol. Biol., Cambridge, United Kingdom |
| Primary Theme and Topics |
Synaptic Transmission and Excitability - Ligand Gated Ion Channels -- Purine receptors |
| Session: |
958. Purine P2X Receptors Poster |
| Presentation Time: | Wednesday, October 27, 2004 2:00 PM-3:00 PM |
| Location: | San Diego Convention Center - Hall A-H, Board # L6 |
| Keywords: | PURINERGIC, GATING, FLUORESCENCE, PERMEABILITY |
The molecular motions that give rise to permeability changes in ATP-gated P2X2 channels are not fully understood (Khakh et al., 1999; Virginio et al., 1999; Eickhorst etal., 2001). We extended work presented previously (Fisher et al., SFN 2003), to show that fusion of the plasma membrane binding pleckstrin homology (PH) domain of phospholipase C to the C termini of P2X2 channels resulted in a splayed conformation of the cytosolic domain, such that the FRET efficiency between CFP and YFP attached to it is reduced from 35±5% to 17±2%. Interestingly, this splayed conformation was not permissive for permeability changes (0.2±0.04 v 2.8±0.4-fold increase in pNMDG+/pNa+ for PH tagged and wt P2X2), or for ATP-evoked changes in FRET measured between fluorophores on the C termini (-1.9±1.8% v –14.1±2.0% change in FY/FC for PH-tagged and wt P2X2). ATP-evoked changes in permeability (2.8±0.8-fold increased pNMDG+/pNa+) and FY/FC (-9.3±1.8% change) were restored for P2X2-PH channels when PIP2 was cleaved to un-tether the PH domain from the plasma membrane, and presumably restore it to a non-splayed conformation. Collectively these data suggest ATP-evoked cytosolic gating motions in P2X2 channels that are associated with permeability changes. If so, how does ATP binding to the extracellular loop cause conformational changes within intracellular domains, when the change has to propagate through the transmembrane segments? In order to address this we have performed alanine-scanning mutagenesis in both the transmembrane domains to identify residues that are involved in conformational changes that give rise to permeability changes. Results from these experiments will be presented along with details of FRET experiments and plausible models of state-specific transmembrane and cytosolic domain gating motions considered.
Supported by MRC, EMBO, HFSP, NIH (GM070925-01)
Sample Citation:
[Authors]. [Abstract Title]. Program No. XXX.XX. 2004 Neuroscience Meeting Planner. San Diego, CA: Society for Neuroscience, 2004. Online.
Copyright © 2004-2025 Society for Neuroscience; all rights reserved. Permission to republish any abstract or part of any abstract in any form must be obtained in writing by SfN office prior to publication.
