<EM>KCNQ2</EM> VARIANT-SPECIFIC SIGNATURES IN CHANNEL FUNCTION, NEURONAL EXCITABILITY AND GENE EXPRESSION

Variants in KCNQ2, a gene encoding a subunit of neuronal voltage-gated K⁺ channels, are linked to benign familial neonatal epilepsy (BFNE), developmental and epileptic encephalopathy (DEE) and intellectual disability (ID). While most studies have examined the effects of KCNQ2 variants on channel function in heterologous systems, their impact on neuronal function remains poorly understood. Here, we assessed the effects of de novo KCNQ2 variants on channel function in CHO cells using patch-clamp electrophysiology and compared them with channel function, neuronal excitability, and gene expression in CRISPR/Cas9-edited human iPSC-derived neurons. In CHO cells, most variants resulted in loss-of-function (LOF) phenotypes impacting ion channel function and/or functional channel density. DEE variants produced more pronounced LOF phenotypes than BFNE variants, whereas the ID variant resulted in a GOF at the level of channel conductance. In neurons, the effects of a BFNE variant resembled those of an acute pharmacological KCNQ-channel block, including decreased neuronal K⁺-current density and action potential frequency. Despite their link to a more severe clinical phenotype, two DEE variants displayed milder or no detectable phenotypes at the level of neuronal currents and excitability. Bulk RNA-sequencing revealed variant-specific changes in gene expression, including synaptic and ion channel-encoding genes. Electrophysiological phenotypes correlated with the number of differentially expressed genes, with heterozygous KCNQ2 mutant neurons exhibiting more pronounced changes than homozygous cultures, suggesting a dominant-negative effect. Together, our data suggest variant-specific phenotypes in KCNQ2-channel function and density that correlate with patient phenotype, which may not be discernible in neurons owing to potential compensatory mechanisms.