A Ca(V)β SH3/Guanylate Kinase Domain Interaction Regulates Multiple Properties of Voltage-gated Ca(2+) Channels

Auxiliary Ca(2+) channel β subunits (Ca(V)β) regulate cellular Ca(2+) signaling by trafficking pore-forming α(1) subunits to the membrane and normalizing channel gating. These effects are mediated through a characteristic src homology 3/guanylate kinase (SH3–GK) structural module, a design feature shared in common with the membrane-associated guanylate kinase (MAGUK) family of scaffold proteins. However, the mechanisms by which the Ca(V)β SH3–GK module regulates multiple Ca(2+) channel functions are not well understood. Here, using a split-domain approach, we investigated the role of the interrelationship between Ca(V)β SH3 and GK domains in defining channel properties. The studies build upon a previously identified split-domain pair that displays a trans SH3–GK interaction, and fully reconstitutes Ca(V)β effects on channel trafficking, activation gating, and increased open probability (P (o)). Here, by varying the precise locations used to separate SH3 and GK domains and monitoring subsequent SH3–GK interactions by fluorescence resonance energy transfer (FRET), we identified a particular split-domain pair that displayed a subtly altered configuration of the trans SH3–GK interaction. Remarkably, this pair discriminated between Ca(V)β trafficking and gating properties: α(1C) targeting to the membrane was fully reconstituted, whereas shifts in activation gating and increased P (o) functions were selectively lost. A more extreme case, in which the trans SH3–GK interaction was selectively ablated, yielded a split-domain pair that could reconstitute neither the trafficking nor gating-modulation functions, even though both moieties could independently engage their respective binding sites on the α(1C) (Ca(V)1.2) subunit. The results reveal that Ca(V)β SH3 and GK domains function codependently to tune Ca(2+) channel trafficking and gating properties, and suggest new paradigms for physiological and therapeutic regulation of Ca(2+) channel activity.