Molecular Determinants of the CaVβ-induced Plasma Membrane Targeting of the CaV1.2 Channel

Ca-V beta subunits modulate cell surface expression and voltage-dependent gating of high voltage-activated (HVA) Ca(V)1 and Ca(V)2 alpha 1 subunits. High affinity Ca-V beta binding onto the so-called alpha interaction domain of the I-II linker of the Ca-V alpha 1 subunit is required for Ca-V beta modulation of HVA channel gating. It has been suggested, however, that Ca-V beta-mediated plasma membrane targeting could be uncoupled from Ca-V beta-mediated modulation of channel gating. In addition to Ca-V beta, Ca-V alpha 2 delta and calmodulin have been proposed to play important roles in HVA channel targeting. Indeed we show that co-expression of Ca-V alpha 2 delta caused a 5-fold stimulation of the whole cell currents measured with Ca(V)1.2 and Ca-V beta 3. To gauge the synergetic role of auxiliary subunits in the steady-state plasma membrane expression of Ca(V)1.2, extracellularly tagged Ca(V)1.2 proteins were quantified using fluorescence-activated cell sorting analysis. Co-expression of Ca(V)1.2 with either Ca-V alpha 2 delta, calmodulin wild type, or apocalmodulin (alone or in combination) failed to promote the detection of fluorescently labeled Ca(V)1.2 subunits. In contrast, co-expression with Ca-V beta 3 stimulated plasma membrane expression of Ca(V)1.2 by a 10-fold factor. Mutations within the alpha interaction domain of Ca(V)1.2 or within the nucleotide kinase domain of Ca-V beta 3 disrupted the Ca-V beta 3-induced plasma membrane targeting of Ca(V)1.2. Altogether, these data support a model where high affinity binding of Ca-V beta to the I-II linker of Ca-V alpha 1 largely accounts for Ca-V beta-induced plasma membrane targeting of Ca(V)1.2.