ADAM17 Mediates Proteolytic Maturation of Voltage-Gated Calcium Channel Auxiliary αδ Subunits, and Enables Calcium Current Enhancement.

The auxiliary αδ subunits of voltage-gated calcium (Ca) channels are key to augmenting expression and function of Ca1 and Ca2 channels, and are also important drug targets in several therapeutic areas, including neuropathic pain. The αδ proteins are translated as preproteins encoding both α and δ, and post-translationally proteolyzed into α and δ subunits, which remain associated as a complex. In this study, we have identified ADAM17 as a key protease involved in proteolytic processing of pro-αδ-1 and αδ-3 subunits.

Biallelic CACNA2D1 loss-of-function variants cause early-onset developmental epileptic encephalopathy.

Voltage-gated calcium (CaV) channels form three subfamilies (CaV1-3). The CaV1 and CaV2 channels are heteromeric, consisting of an α1 pore-forming subunit, associated with auxiliary CaVβ and α2δ subunits. The α2δ subunits are encoded in mammals by four genes, CACNA2D1-4.

Fight or flight: The culprit is lurking in the neighbourhood.

The fight-or-flight response is studied by all students of Physiology as a concerted bodily response to danger. Liu et al (2020) have now revealed its mechanism, after surveying the proteomic neighbourhood around the cardiac calcium channels in a study which is a tour-de-force of modern biological techniques.

Disruption of the Key Ca Binding Site in the Selectivity Filter of Neuronal Voltage-Gated Calcium Channels Inhibits Channel Trafficking.

Voltage-gated calcium channels are exquisitely Ca selective, conferred primarily by four conserved pore-loop glutamate residues contributing to the selectivity filter. There has been little previous work directly measuring whether the trafficking of calcium channels requires their ability to bind Ca in the selectivity filter or to conduct Ca. Here, we examine trafficking of neuronal Ca2.

The αδ-like Protein Cachd1 Increases N-type Calcium Currents and Cell Surface Expression and Competes with αδ-1.

Voltage-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 (Ca2.2) channels, which are important in neurotransmission.

Proteolytic maturation of αδ controls the probability of synaptic vesicular release.

Auxiliary αδ subunits are important proteins for trafficking of voltage-gated calcium channels (Ca) at the active zones of synapses. We have previously shown that the post-translational proteolytic cleavage of αδ is essential for their modulatory effects on the trafficking of N-type (Ca2.2) calcium channels (Kadurin et al.

LRP1 influences trafficking of N-type calcium channels via interaction with the auxiliary αδ-1 subunit.

Voltage-gated Ca (Ca) channels consist of a pore-forming α1 subunit, which determines the main functional and pharmacological attributes of the channel. The Ca1 and Ca2 channels are associated with auxiliary β- and αδ-subunits. The molecular mechanisms involved in αδ subunit trafficking, and the effect of αδ subunits on trafficking calcium channel complexes remain poorly understood.

Proteolytic maturation of αδ represents a checkpoint for activation and neuronal trafficking of latent calcium channels.

The auxiliary αδ subunits of voltage-gated calcium channels are extracellular membrane-associated proteins, which are post-translationally cleaved into disulfide-linked polypeptides α and δ. We now show, using αδ constructs containing artificial cleavage sites, that this processing is an essential step permitting voltage-dependent activation of plasma membrane N-type (Ca2.2) calcium channels.

Thrombospondin-4 reduces binding affinity of [(3)H]-gabapentin to calcium-channel α2δ-1-subunit but does not interact with α2δ-1 on the cell-surface when co-expressed.

The α2δ proteins are auxiliary subunits of voltage-gated calcium channels, and influence their trafficking and biophysical properties. The α2δ ligand gabapentin interacts with α2δ-1, and inhibits calcium channel trafficking. However, α2-1 has also been proposed to play a synaptogenic role, independent of calcium channel function.

Functional exofacially tagged N-type calcium channels elucidate the interaction with auxiliary α2δ-1 subunits.

CaV1 and CaV2 voltage-gated calcium channels are associated with β and α2δ accessory subunits. However, examination of cell surface-associated CaV2 channels has been hampered by the lack of antibodies to cell surface-accessible epitopes and of functional exofacially tagged CaV2 channels. Here we report the development of fully functional CaV2.

The inhibition of functional expression of calcium channels by prion protein demonstrates competition with α2δ for GPI-anchoring pathways.

It has been shown recently that PrP (prion protein) and the calcium channel auxiliary α2δ subunits interact in neurons and expression systems [Senatore, Colleoni, Verderio, Restelli, Morini, Condliffe, Bertani, Mantovani, Canovi, Micotti, Forloni, Dolphin, Matteoli, Gobbi and Chiesa (2012) Neuron 74, 300-313]. In the present study we examined whether there was an effect of PrP on calcium currents. We have shown that when PrP is co-expressed with calcium channels formed from CaV2.

Calcium currents are enhanced by α2δ-1 lacking its membrane anchor.

The accessory α(2)δ subunits of voltage-gated calcium channels are membrane-anchored proteins, which are highly glycosylated, possess multiple disulfide bonds, and are post-translationally cleaved into α(2) and δ. All α(2)δ subunits have a C-terminal hydrophobic, potentially trans-membrane domain and were described as type I transmembrane proteins, but we found evidence that they can be glycosylphosphatidylinositol-anchored. To probe further the function of membrane anchoring in α(2)δ subunits, we have now examined the properties of α(2)δ-1 constructs truncated at their putative glycosylphosphatidylinositol anchor site, located before the C-terminal hydrophobic domain (α(2)δ-1ΔC-term).

Presynaptic HCN1 channels regulate Cav3.2 activity and neurotransmission at select cortical synapses.

The hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are subthreshold, voltage-gated ion channels that are highly expressed in hippocampal and cortical pyramidal cell dendrites, where they are important for regulating synaptic potential integration and plasticity. We found that HCN1 subunits are also localized to the active zone of mature asymmetric synaptic terminals targeting mouse entorhinal cortical layer III pyramidal neurons. HCN channels inhibited glutamate synaptic release by suppressing the activity of low-threshold voltage-gated T-type (Ca(V)3.

A new look at calcium channel α2δ subunits.

The classical roles of α(2)δ proteins are as accessory calcium channel subunits, enhancing channel trafficking. They were thought to have type-I transmembrane topology, but we find that they can form GPI-anchored proteins. Moreover α(2)δ-1 and α(2)δ-3 have been shown to have novel functions in synaptogenesis, independent of their effect on calcium channels.

The alpha2delta subunits of voltage-gated calcium channels form GPI-anchored proteins, a posttranslational modification essential for function.

Voltage-gated calcium channels are thought to exist in the plasma membrane as heteromeric proteins, in which the alpha1 subunit is associated with two auxiliary subunits, the intracellular beta subunit and the alpha(2)delta subunit; both of these subunits influence the trafficking and properties of Ca(V)1 and Ca(V)2 channels. The alpha(2)delta subunits have been described as type I transmembrane proteins, because they have an N-terminal signal peptide and a C-terminal hydrophobic and potentially transmembrane region. However, because they have very short C-terminal cytoplasmic domains, we hypothesized that the alpha(2)delta proteins might be associated with the plasma membrane through a glycosylphosphatidylinositol (GPI) anchor attached to delta rather than a transmembrane domain.

The increased trafficking of the calcium channel subunit alpha2delta-1 to presynaptic terminals in neuropathic pain is inhibited by the alpha2delta ligand pregabalin.

Neuropathic pain results from damage to the peripheral sensory nervous system, which may have a number of causes. The calcium channel subunit alpha(2)delta-1 is upregulated in dorsal root ganglion (DRG) neurons in several animal models of neuropathic pain, and this is causally related to the onset of allodynia, in which a non-noxious stimulus becomes painful. The therapeutic drugs gabapentin and pregabalin (PGB), which are both alpha(2)delta ligands, have antiallodynic effects, but their mechanism of action has remained elusive.

A single conserved proline residue determines the membrane topology of stomatin.

Stomatin is an integral membrane protein which is widely expressed in many cell types. It is accepted that stomatin has a unique hairpin-loop topology: it is anchored to the membrane with an N-terminal hydrophobic domain and the N- and C-termini are cytoplasmically localized. Stomatin is a prototype for a family of related proteins, containing among others MEC-2 (mechanosensory protein 2) from Caenorhabditis elegans, SLP (stomatin-like protein)-3 and podocin, all of which interact with ion channels to regulate their activity.

Differential effects of N-glycans on surface expression suggest structural differences between the acid-sensing ion channel (ASIC) 1a and ASIC1b.

ASICs (acid-sensing ion channels) are H(+)-gated Na(+) channels with a widespread expression pattern in the central and the peripheral nervous system. ASICs have a simple topology with two transmembrane domains, cytoplasmic termini and a large ectodomain between the transmembrane domains; this topology has been confirmed by the crystal structure of chicken ASIC1. ASIC1a and ASIC1b are two variants encoded by the asic1 gene.

Zebrafish acid-sensing ion channel (ASIC) 4, characterization of homo- and heteromeric channels, and identification of regions important for activation by H+.

There are four genes for acid-sensing ion channels (ASICs) in the genome of mammalian species. Whereas ASIC1 to ASIC3 form functional H+-gated Na+ channels, ASIC4 is not gated by H+, and its function is unknown. Zebrafish has two ASIC4 paralogs: zASIC4.

Strong modulation by RFamide neuropeptides of the ASIC1b/3 heteromer in competition with extracellular calcium.

Acid-sensing ion channels are excitatory receptors for extracellular H+. Since the extracellular H+ concentration can significantly increase during an inflammation, one of the proposed functions for ASICs is peripheral perception of pain. The ASIC1b and ASIC3 subunits are specifically expressed in sensory ganglia neurons and are candidate sensors of peripheral acidosis.