Characterization of ATPase Activity of P2RX2 Cation Channel.

Rahul Mittal,M'Hamed Grati,Xi Lin,Miloslav Sedlacek,Xue Z Liu,Denise Yan,Amjad Farooq,Bechara Kachar,Prem Chapagain,Yanbin Zhang,Qing Chang,Fenghua Yuan

doi:10.3389/fphys.2016.00186

Abstract

P2X purinergic receptors are plasma membrane ATP-dependent cation channels that are broadly distributed in the mammalian tissues. P2RX2 is a modulator of auditory sensory hair cell mechanotransduction and plays an important role in hair cell tolerance to noise. In this study, we demonstrate for the first time in vitro and in cochlear neuroepithelium, that P2RX2 possesses the ATPase activity. We observed that the P2RX2 V60L human deafness mutation alters its ability to bind ATP, while the G353R has no effect on ATP binding or hydrolysis. A non-hydrolysable ATP assay using HEK293 cells suggests that ATP hydrolysis plays a significant role in the opening and gating of the P2RX2 ion channel. Moreover, the results of structural modeling of the molecule was in agreement with our experimental observations. These novel findings suggest the intrinsic ATPase activity of P2RX2 and provide molecular insights into the channel opening.

Highlights

P2X receptor family comprises seven different receptors, P2RX1 to P2RX7 (Mittal et al, 2016)
ATP stimulation of HEK293 cells expressing WT P2RX2 evoked a large inward current, as did to a lesser intensity G353R form, but a very faint or no current was obtained with V60L form (Figures 1A,B)
We observed that HEK293 expressing V60L P2RX2 released significantly less inorganic phosphate than WT or G353R P2RX2 (P < 0.001; Figure 2D)

Summary

Introduction

P2X receptor family comprises seven different receptors, P2RX1 to P2RX7 (Mittal et al, 2016). These receptors are expressed in a wide variety of cell types and are involved in numerous physiological processes, including platelet aggregation, immune responses smooth muscle contraction, inflammation, and sensory neurotransmission (Burnstock, 2013; Zhang et al, 2015; Martínez-Ramírez et al, 2016; Riding and Pullar, 2016; Sáez-Orellana et al, 2016) Out of these seven receptors, P2RX2 is an ATP-gated trimeric ion channel that plays an important role in sound transduction and auditory neurotransmission in the inner ear (Housley et al, 2002, 2013; Järlebark et al, 2002; Wang et al, 2003; Yan et al, 2013). PIP ns are involved in the activation of many ion channels and enzymes, and regulate virtually all membrane trafficking events, including

Methods

Results

Conclusion