| We have recently described a novel splice variant of the Cavβ 1 gene, Cavβ1d, which is truncated before the second conserved domain due to alternative splicing. To determine the effects of the β1d subunit on Cav1.2 channels compared to the well-characterized β1b subunit, which modulates gating of channels and promotes trafficking to the surface membrane, we studied HEK 293 cells transiently transfected with the Cav1.2 subunit ± β 1b or β1d subunits. Whole-cell patch clamp studies using 10 mM Ba2+ demonstrated that peak IBa was unchanged comparing α1c alone to α1c + β 1d at all potentials tested, in contrast to 10–12 fold larger currents observed with α1c + β1b. Coexpression of β1b but not β1d with α1c resulted in a leftward shift in the voltage dependence of activation V 1/2 of −3.93 ± .78 for α1c, −4.96 ± 1.06 for α1c + β1b, and −10.19 ± 1.68 for α1c + β1b. Immunocytochemistry and confocal microscopy demonstrated that either α1c alone or α1c + β1d failed to target the channel to the surface membrane, in contrast to coexpression of β1b, which resulted in a punctate pattern of staining, localized to the cell membrane. Results with coexpression of either full-length or partial N-terminal deleted β 1b or β1d showed similar results. Single channel studies with 100 mM Ba2+ at a test potential of –10mV showed that coexpression of both β1btr and β1dtr alter the gating of Cav1.2 channels (Mean Po for α 1c was 0.01 ± 0.002; α1c + β1d = 0.04 ± 0.02; α1c + β1b = 0.05 + 0.01). Together, these results demonstrate that coexpression of the β 1d subunit modulates gating of Cav1.2 Ca2+ channels, but unlike β1b, it fails to target the channel to the membrane. |
