The α2δ Subunit Efficiently Couples VSDs Activation to Pore Opening in Human Cav1.2 Channels

Abstract

The α2δ-1 auxiliary subunit binds to the pore-forming α1c subunit of voltage-gated L-type calcium channels (CaV1.2) and facilitates both membrane trafficking and voltage-dependent activation. Using voltage-clamp fluorometry to study human CaV1.2 channels expressed in Xenopus oocytes, we have recently found that α2δ-1 association, in the presence of β3 subunits, causes voltage sensor domains (VSDs) I, II and III, but not IV, to activate at more hyperpolarized potentials (ΔVhalf= −39mV, −32mV, −18mV for VSDs I-III respectively) and with steeper voltage-dependence (z fold increase with α2δ-1: 1.9, 2.2 and 1.6 for VSDs I-III respectively). Thus, the α2δ-1-induced facilitation of CaV1.2 voltage-dependent activation seems to be mediated through the remodeling of three VSDs.We analyzed our voltage-clamp fluorometry data with a 32-state allosteric model for CaV1.2 activation, consisting of five gating particles (one pore, four VSDs). By simultaneously fitting kinetic and steady-state data from the pore (ionic currents) and from the individual VSDs (fluorescence), we estimated the energetic contribution of each VSD to pore opening in the presence and in the absence of α2δ-1. The model predicts that, in channels not associated with α2δ-1, all VSDs were poorly energetically-coupled to pore opening. The association of α2δ-1 with α1c+β3 specifically increased the coupling energy of VSDs II and III to the pore. In agreement with the model prediction, we found that without α2δ-1 subunits the ionic current deactivation is largely voltage-independent.A low-resolution structure of CaV1.2 channel complexes shows that the α2δ-1 subunit forms a cap that embraces ∼¾ of the extracellular surface of the α1c subunit (Walsh et al., JBC 2009). Based on our results, we propose that the exposed ¼ of the α1c subunit is VSD IV, which does not show evidence of functional interaction with α2δ-1 subunits.