Molecular determinants of the CaVbeta-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 alpha1 subunits. High affinity Ca(V)beta binding onto the so-called alpha interaction domain of the I-II linker of the Ca(V)alpha1 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)alpha2delta and calmodulin have been proposed to play important roles in HVA channel targeting. Indeed we show that co-expression of Ca(V)alpha2delta caused a 5-fold stimulation of the whole cell currents measured with Ca(V)1.2 and Ca(V)beta3. 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)alpha2delta, 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)beta3 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)beta3 disrupted the Ca(V)beta3-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)alpha1 largely accounts for Ca(V)beta-induced plasma membrane targeting of Ca(V)1.2.
Bourdin
Benoîte
B
Institute of Neurophysiology and Center for Molecular Medicine, Cologne University, D-50931 Cologne, Germany.
Marger
Fabrice
F
Wall-Lacelle
Sébastien
S
Schneider
Toni
T
Klein
Hélène
H
Sauvé
Rémy
R
Parent
Lucie
L
D000818:Animals; D019556:COS Cells; D015220:Calcium Channels / Q000737:chemistry / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004553:Electric Conductivity; D006801:Humans; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011817:Rabbits; D051381:Rats
D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D002147:Calmodulin; C419385:L-type calcium channel alpha(1C)