CONNECTION BETWEEN GNB1 ENCEPHALOPATHY-CAUSING MUTATIONS AND M-CURRENT (K V 7.2/7.3)

Voltage-gated potassium channels play an essential role in controlling neuronal excitability. Among them, K_v7.2/7.3 channels, encoded by KCNQ2 and KCNQ3 genes, mediate the M-current, which stabilizes the resting membrane potential and limits repetitive firing. We show that Gβγ subunits regulate K_v7.2/7.3, doubling the currents, affecting the expression and gating of the channel. We investigated whether mutations in the GNB1 gene that encodes Gβ₁, which cause GNB1 encephalopathy, may affect these regulations. We co-expressed K_v7.2/7.3 channels with Gβγ containing wild-type (WT) or mutant Gβ₁ in Xenopus oocytes and used two-electrode voltage clamp to quantify changes in M-current. The G protein-gated channel K_ir3.2 (GIRK2) was used as a control, as the effects of different Gβ₁ mutants on it are well-described. We examined the expression of the channels and Gβγ using Western blot and immunocytochemistry in isolated giant plasma membrane patches. We analyzed six mutations (I80T, I80N, K78R, K89E, L95P, M101V). All mutants except K78R were loss-of-expression, leading to a decrease in Gβγ protein expression and, consequently, a reduction in Gβγ effect on M-current. In contrast, mutation K78R showed gain-of-expression and enhanced the K_v7.2/7.3 current stronger than WT Gβγ. We adjusted RNA concentrations of all mutants to match WT Gβ₁ protein expression levels. Under these conditions, I80T, L96P and M101V regulated K_v7.2/7.3 similarly to WT Gβγ, whereas I80N and K89E mutants demonstrated loss-of-function. These findings identify a novel role for Gβγ in the regulation of neuronal M-current and suggest that impaired Gβγ-K_v7.2/7.3 signaling may contribute to the pathogenesis of neurodevelopmental disorders.