Functional characterization of a novel de novo CACNA1C pathogenic variant in a patient with neurodevelopmental disorder

Abstract

Mutations in CACNA1C, the gene encoding Ca_v1.2 voltage-gated calcium channels, are associated with a spectrum of disorders, including Timothy syndrome and other neurodevelopmental and cardiac conditions. In this study, we report a child with a de novo heterozygous missense variant (c.1973T > C; L658P) in CACNA1C, presenting with refractory epilepsy, global developmental delay, hypotonia, and multiple systemic abnormalities, but without overt cardiac dysfunction. Electrophysiological analysis of the recombinant Ca_v1.2 L658P variant revealed profound gating alterations, most notably a significant hyperpolarizing shift in the voltage dependence of activation and inactivation. Additionally, molecular modeling suggested that the L658P mutation disrupts interactions within the IIS5 transmembrane segment, reducing the energy barrier for state transitions and facilitating channel opening at more negative voltages. These findings establish L658P as a pathogenic CACNA1C variant primarily associated with severe neurological dysfunction and expands the phenotypic spectrum of CACNA1C-related disorders.

Keywords: CACNA1C; Calcium channel; Cav1.2; Channelopathies; Electrophysiology; L658P.

Fig. 1

Electrophysiological properties of the Ca_v1.2 L658P variant. A Location of the L658P missense variant (red dot) within the secondary membrane topology of Ca_v1.2 channel. B Amino acid sequence alignment of the IIS5 transmembrane segment showing the conservation of the L658 residue across the 10 human voltage-gated calcium channel isoforms. Alignments were performed using UniProt (Ca_v1.1 Q13698; Ca_v1.2 Q13936; Ca_v1.3 Q01668; Ca_v1.4 O60840; Ca_v2.1 O00555; Ca_v2.2 Q00975; Ca_v2.3 Q15878; Ca_v3.1 O43497; Ca_v3.2 O95180; Ca_v3.3 Q9P0X4). C Representative L-type current traces recorded from tsA-201 cells expressing Ca_v1.2 wild-type (WT, black traces) and L658P variant channels (red traces), in combination with Ca_vβ₂, and Ca_vα₂δ₁. D Corresponding mean current–voltage (I/V) relationship. E Mean maximal macroscopic conductance (G_max) obtained from the fit of the I/V curves with the modified Boltzmann Eq. (1). F Mean reversal potential (V_rev) values. G Mean normalized voltage dependence of activation. H Mean half-activation potential (V_0.5 activation) values obtained from the fit of the activation curves with the modified Boltzmann Eq. (2). I Mean normalized voltage dependence of steady state inactivation. J Mean half-inactivation potential (V_0.5 inactivation) values obtained from the fit of the inactivation curves with the modified Boltzmann Eq. (3). K Mean non-inactivating channel fraction values. L Mean window currents calculated from the activation and inactivation curves using the Eq. (4). The squared area shows the window current between − 70 and − 55 mV, corresponding to a range of neuronal resting membrane potentials. M Mean window current values calculated from the area under the curve between − 70 and − 55 mV. N Mean time constant τ values of current activation kinetics. O Mean time constant τ values of current inactivation kinetics. P Mean normalized recovery of inactivation kinetics. Q Mean time constant τ values of recovery from inactivation obtained by fitting the recovery curves with a single-exponential Eq. (5). R Dose–response of Ca_v1.2 channels for isradipine. S Dose–response of Ca_v1.2 channels for verapamil. T Molecular modeling of Ca_v1.2 showing the impact of the L658P mutation on various superimposed Ca_v1.2 channel states including resting (cyan), near-open (green), and high-voltage inactivated states (yellow and orange)