Alternative splicing of the voltage-gated Ca²⁺ channel β₄ subunit creates a uniquely folded N-terminal protein binding domain with cell-specific expression in the cerebellar cortex

Ca²⁺ β channel subunits regulate cell-surface expression and gating of voltage-dependent Ca²⁺ channel α1 subunits. Based on primary sequence comparisons, β subunits are predicted to be modular structures composed of five domains (A-E) that are related to the large family of membrane-associated guanylate kinase proteins. The crystal structure of the β subunit core B-D domains has been reported recently; however, little is known about the structures of the A and E domains. The N-terminal A domain differs among the four subtypes of Ca²⁺ channel β subunits (β₁-β₄) primarily as the result of two duplications of an ancestral gene containing multiple alternatively spliced exons. At least nine A domain sequences can be generated by alternative splicing. In this report, we focus on one A domain sequence, the highly conserved β_4a A domain. We solved its three-dimensional structure and show that it is expressed in punctate structures throughout the molecular layer of the cerebellar cortex. We also demonstrate that it does not participate directly in Ca_v2.1 Ca²⁺ channel gating but serves as a binding site in protein-protein interactions with synaptotagmin I and the LC2 domain of microtubule-associated protein 1A. With respect to β₄ subunits, the interactions are specific for the β_4a splice variant, because they do not occur with the β_4b A domain. These results have strong bearing on our current understanding of the structure of alternatively spliced Ca²⁺ channel β subunits and the cell-specific roles they play in the CNS.