Expression and Modulation of an Invertebrate Presynaptic Calcium Channel α1 Subunit Homolog

J David Spafford,Lina Chen,Zhong-Ping Feng,August B Smit,Gerald W Zamponi

doi:10.1074/jbc.m302212200

J David Spafford, Lina Chen + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m302212200

Copy DOI

Abstract

Here we report the first assessment of the expression and modulation of an invertebrate alpha1 subunit homolog of mammalian presynaptic Cav2 calcium channels (N-type and P/Q-type) in mammalian cells. Our data show that molluscan channel (LCav2a) isolated from Lymnaea stagnalis is effectively membrane-targeted and electrophysiologically recordable in tsA-201 cells only when the first 44 amino acids of LCav2a are substituted for the corresponding region of rat Cav2.1. When coexpressed with rat accessory subunits, the biophysical properties of LCav2a-5'rbA resemble those of mammalian N-type calcium channels with respect to activation and inactivation, lack of pronounced calcium dependent inactivation, preferential permeation of barium ions, and cadmium block. Consistent with reports of native Lymnaea calcium currents, the LCav2a-5'rbA channel is insensitive to micromolar concentrations of omega-conotoxin GVIA and is not affected by nifedipine, thus confirming that it is not of the L-type. Interestingly, the LCav2a-5'rbA channel is almost completely and irreversibly inhibited by guanosine 5'-3-O-(thio)triphosphate but not regulated by syntaxin1, suggesting that invertebrate presynaptic calcium channels are differently modulated from their vertebrate counterparts.

Highlights

We report the first assessment of the expression and modulation of an invertebrate ␣1 subunit homolog of mammalian presynaptic Cav2 calcium channels (Ntype and P/Q-type) in mammalian cells
This possibly serves to optimize the efficiency of synaptic transmission and as a negative feedback mechanism allowing the regulation of calcium channel activity during various steps of exocytosis [7, 12]
Our data revealed that synaptic transmission in identified Lymnaea neurons was dependent on a Cav2 calcium channel homolog (LCav2a) that is capable of associating with the scaffolding proteins Mint-1 and CASK but which curiously lacks the synaptic protein interaction site common to mammalian presynaptic calcium channels [21]

Summary

Introduction

We report the first assessment of the expression and modulation of an invertebrate ␣1 subunit homolog of mammalian presynaptic Cav2 calcium channels (Ntype and P/Q-type) in mammalian cells. We attempted a strategy used previously to obtain expression of a mammalian Cav2.2 calcium channel variant in Xenopus oocytes [27,28,29] by replacing part of the N-terminal region of the LCav2a channel (44 amino acids) with the corresponding sequence from rat Cav2.1 (creating LCav2a-5ЈrbA).

Results

Conclusion