Role of a conserved glutamine in the function of voltage-gated Ca2+ channels revealed by a mutation in human CACNA1D

Voltage-gated Ca v Ca 2+ channels play crucial roles in regulating gene transcription, neuronal excitability, and synaptic transmission. Natural or pathological variations in Ca v channels have yielded rich insights into the molecular determinants controlling channel function. Here, we report the consequences of a natural, putatively disease-associated mutation in the CACNA1D gene encoding the pore-forming Ca v 1.3 α 1 subunit. The mutation causes a substitution of a glutamine residue that is highly conserved in the extracellular S1–S2 loop of domain II in all Ca v channels with a histidine and was identified by whole-exome sequencing of an individual with moderate hearing impairment, developmental delay, and epilepsy. When introduced into the rat Ca v 1.3 cDNA, Q558H significantly decreased the density of Ca 2+ currents in transfected HEK293T cells. Gating current analyses and cell-surface biotinylation experiments suggested that the smaller current amplitudes caused by Q558H were because of decreased numbers of functional Ca v 1.3 channels at the cell surface. The substitution also produced more sustained Ca 2+ currents by weakening voltage-dependent inactivation. When inserted into the corresponding locus of Ca v 2.1, the substitution had similar effects as in Ca v 1.3. However, the substitution introduced in Ca v 3.1 reduced current density, but had no effects on voltage-dependent inactivation. Our results reveal a critical extracellular determinant of current density for all Ca v family members and of voltage-dependent inactivation of Ca v 1.3 and Ca v 2.1 channels.