Identification of P2X2/P2X4/P2X6 heterotrimeric receptors using atomic force microscopy (AFM) imaging

Author(s)

Antonio, Ligia S. ^{[1, 2]} ; Stewart, Andrew P. ^[1] ; Varanda, Wamberto A. ^[2] ; Edwardson, J. Michael ^[1]

Affiliation

^[1] Univ Cambridge, Dept Pharmacol, Cambridge CB2 1PD - England

^[2] Univ Sao Paulo, Dept Physiol, Sch Med Ribeirao Preto, BR-14049 Ribeirao Preto, SP - Brazil

• Document type: Journal article
• Source: FEBS Letters; v. 588, n. 12, p. 2125-2128, JUN 5 2014.
• Web of Science Citations: 7

Abstract

Seven P2X purinergic receptor subunits have been identified: P2X1-P2X7. The overlapping expression of P2X2, P2X4 and P2X6 subunits has been shown in different cell types, and functional analysis of P2X receptors in Leydig cells suggests that the three subunits might interact. Here, His(6)-tagged P2X2, HA-tagged P2X4 and FLAG-tagged P2X6 subunits were co-expressed in tsA 201 cells. After sequential co-immunoprecipitation using anti-HA and anti-FLAG beads, all three subunits were present, demonstrating their interaction. Atomic force microscopy (AFM) imaging revealed receptors that were specifically decorated by both an anti-His(6) antibody and an anti-HA Fab fragment, indicating the presence of a P2X2/4/6 heterotrimer. To our knowledge, this is the first report of a P2X receptor containing three different subunits. (C) 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved. (AU)

• FAPESP's process: 08/10000-0 - P2X subunits expression in mice Leydig cells at different ages and interactions amongst P2X2, P2X4, P2X6 subunits forming functional purinergic receptors
• Grantee: Ligia Subitoni Antonio
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 06/50954-7 - Ionic currents and receptors in the physiology of excitable and non-excitable cells
• Grantee: Wamberto Antonio Varanda
• Support Opportunities: Research Projects - Thematic Grants