The Calcium Channel α2δ-2 Subunit Partitions with CaV2.1 into Lipid Rafts in Cerebellum: Implications for Localization and Function

By A. Davies, L. Douglas, J. Hendrich, J. Wratten, Alexandra Tran Van Minh, I. Foucault, Dietlind Koch, W. S. Pratt, H. Saibil, A. Dolphin. Journal of Neuroscience, 2006

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The accessory α2δ subunits of voltage-gated calcium channels are highly glycosylated transmembrane proteins that interact with calcium channel α1 subunits to enhance calcium currents. We compared the membrane localization and processing of native cerebellar α2δ-2 subunits with α2δ-2 stably expressed in tsA-201 cells. We identified that α2δ-2 is completely concentrated in cholesterol-rich microdomains (lipid rafts) in cerebellum, in which it substantially colocalizes with the calcium channel α1 subunit CaV2.1, although CaV2.1 is also present in the Triton X-100-soluble fraction. In tsA-201 cells, unlike cerebellum, α2δ-2 is not completely proteolytically processed into α2-2 and δ-2. However, this processing is more complete in the lipid raft fraction of tsA-201 cells, in which α2δ-2 also colocalizes with CaV2.1. Cholesterol depletion of intact cells disrupted their lipid rafts and enhanced CaV2.1/α2δ-2/β4 currents. Furthermore, α2δ-2 coimmunoprecipitates with lipid raft-associated proteins of the stomatin family. The apparent affinity of α2δ-2 for its ligand gabapentin is increased markedly in the cholesterol-rich microdomain fractions, in both cerebellum and the stable α2δ-2 cell line. In contrast, α2δ-2 containing a point mutation (R282A) has a much lower affinity for gabapentin, and this is not enhanced in the lipid raft fraction. This R282A mutant α2δ-2 shows reduced functionality in terms of enhancement of CaV2.1/β4 calcium currents, suggesting that the integrity of the gabapentin binding site may be important for normal functioning of α2δ-2. Together, these results indicate that both α2δ-2 and CaV2.1 are normally associated with cholesterol-rich microdomains, and this influences their functionality.

Subfields: Cell Biology, Cellular and Molecular Neuroscience, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/c74b728045334db89b82b7638d2f58c4979f31bb

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