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Molecular determinants of the CaVbeta-induced plasma membrane targeting of the CaV1.2 channel

Benoîte Bourdin1, Fabrice Marger, Sébastien Wall-Lacelle

  • 1Institute of Neurophysiology and Center for Molecular Medicine, Cologne University, D-50931 Cologne, Germany.

The Journal of Biological Chemistry
|May 19, 2010
PubMed

Insights

Calcium channel beta (Ca(V)beta) subunits are crucial for targeting Ca(V)1.2 channels to the cell surface. High-affinity binding of Ca(V)beta to the Ca(V)alpha1 subunit

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biophysics

Background:

  • Calcium channel auxiliary subunits, including Ca(V)beta, Ca(V)alpha2delta, and calmodulin, are known to modulate the function and trafficking of high voltage-activated (HVA) calcium channels.
  • The precise mechanisms by which these subunits influence the plasma membrane expression of Ca(V)1.2 channels are not fully elucidated.
  • While Ca(V)beta binding to the I-II linker of Ca(V)alpha1 is essential for gating modulation, its role in plasma membrane targeting has been debated.

Purpose of the Study:

  • To investigate the role of Ca(V)beta subunits in the plasma membrane targeting of Ca(V)1.2 channels.
  • To determine whether Ca(V)beta-mediated plasma membrane targeting can be dissociated from Ca(V)beta-mediated gating modulation.
  • To compare the effects of Ca(V)beta subunits with those of Ca(V)alpha2delta and calmodulin on Ca(V)1.2 trafficking.

Main Methods:

  • Co-expression of Ca(V)1.2 with various auxiliary subunits (Ca(V)beta3, Ca(V)alpha2delta, calmodulin) in a cellular system.
  • Quantification of whole-cell currents using electrophysiology to assess channel activity.
  • Utilized extracellularly tagged Ca(V)1.2 proteins and fluorescence-activated cell sorting (FACS) analysis to measure steady-state plasma membrane expression.
  • Introduced mutations in the Ca(V)alpha1 I-II linker and Ca(V)beta3 nucleotide kinase domain to probe interaction sites.

Main Results:

  • Co-expression of Ca(V)alpha2delta alone resulted in a 5-fold increase in Ca(V)1.2 currents but did not significantly enhance plasma membrane expression.
  • Ca(V)beta3 significantly stimulated Ca(V)1.2 plasma membrane expression by a 10-fold factor, as measured by FACS.
  • Mutations disrupting the high-affinity interaction between Ca(V)beta3 and the Ca(V)alpha1 I-II linker abolished the Ca(V)beta3-induced plasma membrane targeting.
  • Ca(V)alpha2delta and calmodulin did not promote Ca(V)1.2 plasma membrane targeting, either alone or in combination.

Conclusions:

  • High-affinity binding of Ca(V)beta subunits to the I-II linker of Ca(V)alpha1 is a critical determinant for the plasma membrane targeting of Ca(V)1.2 channels.
  • Ca(V)beta-mediated plasma membrane targeting of Ca(V)1.2 is largely dependent on this specific interaction.
  • The findings support a model where Ca(V)beta plays a dominant role in trafficking Ca(V)1.2 channels to the cell surface, distinct from the roles of Ca(V)alpha2delta and calmodulin in this process.

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