Determinants of the voltage dependence of G protein modulation within calcium channel β subunits

Ca-V beta subunits of voltage-gated calcium channels contain two conserved domains, a src-homology-3 (SH3) domain and a guanylate kinase-like (GK) domain with an intervening HOOK domain. We have shown in a previous study that, although G beta gamma-mediated inhibitory modulation of Ca(V)2.2 channels did not require the interaction of a Ca-V beta subunit with the Ca-V alpha 1 subunit, when such interaction was prevented by a mutation in the alpha 1 subunit, G protein modulation could not be removed by a large depolarization and showed voltage-independent properties (Leroy et al., J Neurosci 25:6984-6996, 2005). In this study, we have investigated the ability of mutant and truncated Ca-V beta subunits to support voltage-dependent G protein modulation in order to determine the minimal domain of the Ca-V beta subunit that is required for this process. We have coexpressed the Ca-V beta subunit constructs with Ca(V)2.2 and alpha(2)delta-2, studied modulation by the activation of the dopamine D2 receptor, and also examined basal tonic modulation. Our main finding is that the Ca-V beta subunit GK domains, from either beta 1b or beta 2, are sufficient to restore voltage dependence to G protein modulation. We also found that the removal of the variable HOOK region from beta 2a promotes tonic voltage-dependent G protein modulation. We propose that the absence of the HOOK region enhances G beta gamma binding affinity, leading to greater tonic modulation by basal levels of G beta gamma. This tonic modulation requires the presence of an SH3 domain, as tonic modulation is not supported by any of the Ca-V beta subunit GK domains alone.