title: Reciprocal interactions regulate targeting of calcium channel beta subunits and membrane expression of alpha1 subunits in cultured hippocampal neurons.
abstract: Auxiliary beta subunits modulate current properties and mediate the functional membrane expression of voltage-gated Ca(2+) channels in heterologous cells. In brain, all four beta isoforms are widely expressed, yet little is known about their specific roles in neuronal functions. Here, we investigated the expression and targeting properties of beta subunits and their role in membrane expression of Ca(V)1.2 alpha(1) subunits in cultured hippocampal neurons. Quantitative reverse transcription-PCR showed equal expression, and immunofluorescence showed a similar distribution of all endogenous beta subunits throughout dendrites and axons. High resolution microscopy of hippocampal neurons transfected with six different V5 epitope-tagged beta subunits demonstrated that all beta subunits were able to accumulate in synaptic terminals and to colocalize with postsynaptic Ca(V)1.2, thus indicating a great promiscuity in alpha(1)-beta interactions. In contrast, restricted axonal targeting of beta(1) and weak colocalization of beta(4b) with Ca(V)1.2 indicated isoform-specific differences in local channel complex formation. Membrane expression of external hemagglutinin epitope-tagged Ca(V)1.2 was strongly enhanced by all beta subunits in an isoform-specific manner. Conversely, mutating the alpha-interaction domain of Ca(V)1.2 (W440A) abolished membrane expression and targeting into dendritic spines. This demonstrates that in neurons the interaction of a beta subunit with the alpha-interaction domain is absolutely essential for membrane expression of alpha(1) subunits, as well as for the subcellular localization of beta subunits, which by themselves possess little or no targeting properties.
authors.affiliation: Department of Physiology and Medical Physics, Innsbruck Medical University, Austria. Gerald.Obermair@i-med.ac.at
authors.forename: Gerald J
authors.initials: GJ
authors.lastname: Obermair
authors.forename: Bettina
authors.initials: B
authors.lastname: Schlick
authors.forename: Valentina
authors.initials: V
authors.lastname: Di Biase
authors.forename: Prakash
authors.initials: P
authors.lastname: Subramanyam
authors.forename: Mathias
authors.initials: M
authors.lastname: Gebhart
authors.forename: Sabine
authors.initials: S
authors.lastname: Baumgartner
authors.forename: Bernhard E
authors.initials: BE
authors.lastname: Flucher
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism; D006624:Hippocampus / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D020125:Mutation, Missense; D017729:Presynaptic Terminals / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000502:physiology; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: C514350:CACNA1C protein, mouse; D020746:Calcium Channels, L-Type; D021122:Protein Subunits