Phosphatidylinositol 4,5-Bisphosphate Rescues TRPM4 Channels from Desensitization

Haruhisa Okawa,Yuanyuan Wang,Zheng Zhang,Emily R Liman

doi:10.1074/jbc.m506965200

Haruhisa Okawa, Yuanyuan Wang + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m506965200

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Abstract

TRPM4 is a Ca(2+)-activated nonselective cation channel that regulates membrane potential in response to intracellular Ca(2+) signaling. In lymphocytes it plays an essential role in shaping the pattern of intracellular Ca(2+) oscillations that lead to cytokine secretion. To better understand its role in this and other physiological processes, we investigated mechanisms by which TRPM4 is regulated. TRPM4 was expressed in ChoK1 cells, and currents were measured in excised patches. Under these conditions, TRPM4 currents were activated by micromolar concentrations of cytoplasmic Ca(2+) and progressively desensitized. Here we show that desensitization can be explained by a loss of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) from the channels. Poly-l-lysine, a PI(4,5)P(2) scavenger, caused rapid desensitization, whereas MgATP, at concentrations that activate lipid kinases, promoted recovery of TRPM4 currents. Application of exogenous PI(4,5)P(2) to the intracellular surface of the patch restored the properties of TRPM4 currents. Our results suggest that PI(4,5)P(2) acts to uncouple channel opening from changes in the transmembrane potential, allowing current activation at physiological voltages. These data argue that hydrolysis of PI(4,5)P(2) underlies desensitization of TRPM4 and support the idea that PI(4,5)P(2) is a general regulator for the gating of TRPM ion channels.

Highlights

We found that TRPM5 channels desensitize to Ca2ϩ and that sensitivity could be recovered by exogenous PI[4,5]P2 [6]
Recovery of TRPM4 Currents from Rundown by MgATP—To study second messenger regulation of TRPM4 channels, we expressed mouse TRPM4 fused to cyan fluorescent protein in ChoK1 cells, which otherwise show little endogenous TRPM4-like channel activity
Consistent with previous reports (e.g. Ref. 24), in excised inside-out patches, TRPM4 currents could be evoked in response to 100 ␮M cytosolic Ca2ϩ, and this Ca2ϩ-evoked current ran down over several seconds (Fig. 1A)

Summary

Introduction

It has been proposed to play a role in myogenic constriction of cerebral arteries [10] Both TRPM4 and TRPM5 appear to be activated downstream of phospholipase C (PLC) signaling, most likely through the direct effects of store-released Ca2ϩ on channel gating (3, 6 – 8). The sensitivity of TRPM4 channels to Ca2ϩ changes subsequent to activation [22], which may serve a physiologically important role in allowing the channels to respond over a greater stimulus range. We find that manipulations that are expected to change endogenous PI[4,5]P2 levels predictably enhance or diminish desensitization of TRPM4 Together these data indicate that hydrolysis of PI[4,5]P2 underlies desensitization of TRPM4

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