Stargazin Modulates Neuronal Voltage-dependent Ca2+ Channel Cav2.2 by a Gβγ-dependent Mechanism

Isabella Tselnicker,Vladimir A Tsemakhovich,Carmen W Dessauer,Nathan Dascal

doi:10.1074/jbc.m110.121277

Abstract

Loss of neuronal protein stargazin (gamma(2)) is associated with recurrent epileptic seizures and ataxia in mice. Initially, due to homology to the skeletal muscle calcium channel gamma(1) subunit, stargazin and other family members (gamma(3-8)) were classified as gamma subunits of neuronal voltage-gated calcium channels (such as Ca(V)2.1-Ca(V)2.3). Here, we report that stargazin interferes with G protein modulation of Ca(V)2.2 (N-type) channels expressed in Xenopus oocytes. Stargazin counteracted the Gbetagamma-induced inhibition of Ca(V)2.2 channel currents, caused either by coexpression of the Gbetagamma dimer or by activation of a G protein-coupled receptor. Expression of high doses of Gbetagamma overcame the effects of stargazin. High affinity Gbetagamma scavenger proteins m-cbetaARK and m-phosducin produced effects similar to stargazin. The effects of stargazin and m-cbetaARK were not additive, suggesting a common mechanism of action, and generally independent of the presence of the Ca(V)beta(3) subunit. However, in some cases, coexpression of Ca(V)beta(3) blunted the modulation by stargazin. Finally, the Gbetagamma-opposing action of stargazin was not unique to Ca(V)2.2, as stargazin also inhibited the Gbetagamma-mediated activation of the G protein-activated K(+) channel. Purified cytosolic C-terminal part of stargazin bound Gbetagamma in vitro. Our results suggest that the regulation by stargazin of biophysical properties of Ca(V)2.2 are not exerted by direct modulation of the channel but via a Gbetagamma-dependent mechanism.

Highlights

Stargazin (␥2) is a neuronal protein belonging to a family of transmembrane AMPA3 receptor regulatory proteins, whose mutations are associated with recurrent epileptic seizures and ataxic gait in mice [1]
Binding of G␤␥ to the ␣1 subunit of CaV2.2 produces an array of biophysical effects, including a decrease in current amplitude, which can be relieved by strong depolarization; slowing of the kinetics of activation and inactivation; and a depolarizing shift of voltage dependences of activation and inactivation
We propose that stargazin is not a genuine subunit of Ca2ϩ channels; it regulates CaV2 and probably other G␤␥ effectors by opposing G␤␥-mediated effects

Summary

Introduction

Stargazin (␥2) is a neuronal protein belonging to a family of transmembrane AMPA3 receptor regulatory proteins, whose mutations are associated with recurrent epileptic seizures and ataxic gait in mice [1]. We report that stargazin interferes with G protein modulation of CaV2.2 (N-type) channels expressed in Xenopus oocytes. Stargazin counteracted the G␤␥-induced inhibition of CaV2.2 channel currents, caused either by coexpression of the G␤␥ dimer or by activation of a G protein-coupled receptor.

Results

Conclusion