Native TRPC7 Channel Activation by an Inositol Trisphosphate Receptor-dependent Mechanism

Yasuo Mori,Guillermo Vazquez,James W Putney,Gary St J Bird

doi:10.1074/jbc.m604994200

Yasuo Mori, Guillermo Vazquez + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m604994200

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Abstract

In DT40 B lymphocytes, Canonical Transient Receptor Potential 7 (TRPC7) functions as a diacylglycerol-activated non-selective cation channel. However, previous work indicated that the non-store-operated Ca2+ entry in this cell type depends upon inositol trisphosphate receptors (IP3R). With the cell-attached configuration oleyl-acetyl-glycerol (OAG) induced single channel activity (75 pS) that was not observed in TRPC7-/- cells but was rescued by expression of TRPC7 under conditions expected to produce relatively low levels of expression ((LowT7)TRPC7-/-). A DT40 cell line lacking IP3R(IP3R-/- cells) showed no OAG-induced single channel activity, but this activity was rescued by transient expression of an IP3R((IP3R)IP3R-/-). Single channel properties in (LowT7)TRPC7-/- or (IP3R)IP3R-/- DT40 cells were indistinguishable from one another and from wild-type cells. Thus, TRPC7 forms, or is part of, the channel underlying endogenous diacylglycerol-activated currents in DT40 B lymphocytes, and this activity of native TRPC7 requires IP3R. However, with conditions expected to produce greater expression levels, TRPC7 functioned independently of the presence of IP3R. This finding may serve to resolve previously conflicting reports from expression studies of TRPC channels.

Highlights

Canonical Transient Receptor Potential (TRPC)2 channels, which belong to the larger superfamily of mammalian TRP channel-forming proteins, are among the most important neurotransmitter and hormone-regulated cation channels in nonexcitable cells [1, 2]
In a recent study [7] we reported responses to the synthetic DAG analog oleyl-acetyl-glycerol (OAG) in wild-type DT40 B lymphocytes by using conditions that provide strong inhibition of protein kinase C (PKC)
Stimulation and, of significance, both phospholipase C (PLC)- and OAG-dependent whole cell currents were absent in TRPC7 knock-out (TRPC7Ϫ/Ϫ) DT40 cells, suggesting that the DAG-sensitive activity is mediated by channels containing native, endogenously expressed TRPC7

Summary

Introduction

Canonical Transient Receptor Potential (TRPC) channels, which belong to the larger superfamily of mammalian TRP channel-forming proteins, are among the most important neurotransmitter and hormone-regulated cation channels in nonexcitable cells [1, 2]. TRPC channels function as multifunctional calcium-permeable cation channels that can be activated through the phospholipase C (PLC) pathway. In the avian B-cell line, TRPC7 appears to function as a PLC-regulated, DAG-activated non-selective cation channel. Previous studies from this laboratory [8] and others [9, 10] showed that in DT40 cells non-store-operated Ca2ϩ, receptor-regulated cation entry depends in some manner on inositol trisphosphate (IP3) receptors, but the precise role of IP3 receptors (IP3R) in this pathway has not been defined. We further explored the role of native TRPC7 and IP3R in DT40 cells by characterizing the single channel properties of the channels underlying non-store-operated, DAG-activated currents and evaluating its dependence on IP3R expression

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