Reciprocal interactions regulate targeting of calcium channel β subunits and membrane expression of α1 subunits in cultured hippocampal neurons

Auxiliary β subunits modulate current properties and mediate the functional membrane expression of voltage-gated Ca2+ channels in heterologous cells. In brain, all four β isoforms are widely expressed, yet little is known about their specific roles in neuronal functions. Here, we investigated the expression and targeting properties of β subunits and their role in membrane expression of CaV1.2α1 subunits in cultured hippocampal neurons. Quantitative reverse transcription-PCR showed equal expression, and immunofluorescence showed a similar distribution of all endogenous β subunits throughout dendrites and axons. High resolution microscopy of hippocampal neurons transfected with six different V5 epitope-tagged β subunits demonstrated that all β subunits were able to accumulate in synaptic terminals and to colocalize with postsynaptic Ca V1.2, thus indicating a great promiscuity in α1- β interactions. In contrast, restricted axonal targeting of β1 and weak colocalization of β4b with Ca V1.2 indicated isoform-specific differences in local channel complex formation. Membrane expression of external hemagglutinin epitope-tagged Ca V1.2 was strongly enhanced by all β subunits in an isoform-specific manner. Conversely, mutating the α-interaction domain of CaV1.2 (W440A) abolished membrane expression and targeting into dendritic spines. This demonstrates that in neurons the interaction of a β subunit with the α-interaction domain is absolutely essential for membrane expression of α1 subunits, as well as for the subcellular localization of β subunits, which by themselves possess little or no targeting properties. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.