Abstract
SummaryVoltage-gated calcium channel auxiliary α2δ subunits are important for channel trafficking and function. Here, we compare the effects of α2δ-1 and an α2δ-like protein called Cachd1 on neuronal N-type (CaV2.2) channels, which are important in neurotransmission. Previous structural studies show the α2δ-1 VWA domain interacting with the first loop in CaV1.1 domain-I via its metal ion-dependent adhesion site (MIDAS) motif and additional Cache domain interactions. Cachd1 has a disrupted MIDAS motif. However, Cachd1 increases CaV2.2 currents substantially (although less than α2δ-1) and increases CaV2.2 cell surface expression by reducing endocytosis. Although the effects of α2δ-1 are abolished by mutation of Asp122 in CaV2.2 domain-I, which mediates interaction with its VWA domain, the Cachd1 responses are unaffected. Furthermore, Cachd1 co-immunoprecipitates with CaV2.2 and inhibits co-immunoprecipitation of α2δ-1 by CaV2.2. Cachd1 also competes with α2δ-1 for effects on trafficking. Thus, Cachd1 influences both CaV2.2 trafficking and function and can inhibit responses to α2δ-1.
Highlights
Voltage-gated calcium (CaV) channels are key constituents of excitable cells, including muscles, neurons, and secretory cells, and are essential for their function
By mutating the metal ion-dependent adhesion site (MIDAS) motif in the von Willebrand factor A (VWA) domain of a2d subunits, we have shown that the VWA domains of both a2d-1 and a2d-2 are key to promoting calcium channel trafficking and function (Cantıet al., 2005; Cassidy et al, 2014; Hoppa et al, 2012)
Disruption of the Interaction Site between CaV2.2 and the a2d-1 VWA Domain Prevents the Interaction between a2d-1 and CaV2.2 In previous studies, we found that mutation of the MIDAS motif in a2d-1 and a2d-2 prevented the ability of these proteins to traffic CaV2 channels and abolished the increase in CaV1 and CaV2 currents, normally seen with wild-type (WT) a2d-1 and a2d-2 (Cantı et al, 2005; Cassidy et al, 2014; Hoppa et al, 2012)
Summary
Voltage-gated calcium (CaV) channels are key constituents of excitable cells, including muscles, neurons, and secretory cells, and are essential for their function (for a review, see Zamponi et al, 2015). CaV a1 subunits form the pore of the channels, determining their main biophysical and pharmacological properties (Zamponi et al, 2015), but the associated b and a2d proteins represent auxiliary subunits that are important contributors to the trafficking and biophysical properties of the channel complexes (Gurnett et al, 1996; Leung et al, 1987; Pragnell et al, 1994; Takahashi et al, 1987). The mechanism by which the a2d subunits increase trafficking and function of channel complexes is less well understood (Cantıet al., 2005; Cassidy et al, 2014; Ferron et al, 2018; Kadurin et al, 2016; Savalli et al, 2016). The a2d proteins undergo several post-translational processing steps, including N-glycosylation, proteolytic cleavage into a2 and d (De Jongh et al, 1990; Ellis et al, 1988; Jay et al, 1991), and glycosyl-phosphatidylinositol (GPI) anchoring (Davies et al, 2010)
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