Voltage-gated rearrangements associated with differential β-subunit modulation of the L-type Ca²⁺ channel inactivation

Auxiliary β-subunits bound to the cytoplasmic α₁- interaction domain of the pore-forming α_1C-subunit are important modulators of voltage-gated Ca²⁺ channels. The underlying mechanisms are not yet well understood. We investigated correlations between differential modulation of inactivation by β_1a- and β₂-subunits and structural responses of the channel to transition into distinct functional states. The NH₂-termini of the α_1C- and β-subunits were fused with cyan or yellow fluorescent proteins, and functionally coexpressed in COS1 cells. Fluorescence resonance energy transfer (FRET) between them or with membrane-trapped probes was measured in live cells under voltage clamp. It was found that in the resting state, the tagged NH₂-termini of the α_1C- and β-subunit fluorophores are separated. Voltage-dependent inactivation generates strong FRET between α_1C and β_1a suggesting mutual reorientation of the NH₂-termini, but their distance vis-à-vis the plasma membrane is not appreciably changed. These voltage-gated rearrangements were substantially reduced when the β_1a-subunit was replaced by β₂. Differential β-subunit modulation of inactivation and of FRET between α_1C and β were eliminated by inhibition of the slow inactivation. Thus, differential β-subunit modulation of inactivation correlates with the voltage-gated motion between the NH₂-termini of α_1C- and β-subunits and targets the mechanism of slow voltage-dependent inactivation.