To understand how various splicing factors regulate alternative splicing of voltage-gated potassium channels Kv7.2 and Kv3.3.

Voltage-gated potassium channels (VGKCs) are widely expressed across neurons, muscle, and cardiac tissue, playing pivotal roles in essential physiological functions. In humans, the VGKC subunit Kv7.2, encoded by the KCNQ2 gene, is highly expressed in the cerebral cortex and cerebellum, while Kv3.3, encoded by the KCNC3 gene, is highly expressed in the cerebellum and basal ganglia. Kv7.2 mutations are linked to benign familial neonatal convulsions (BFNC), while Kv3.3 mutations are associated with spinocerebellar ataxia type-13 (SCA-13). Multiple disease-causing mutations in Kv7.2 and Kv3.3, including alternative splice site mutations, cluster in the long cytoplasmic C-terminal domain. This region is crucial for channel function and regulation, as it contains domains essential for subunit assembly and for a complex network of mutually interacting regulatory molecules such as calmodulin, PIP2, syntaxin-1A, and protein kinase C-regulated factors. At least 10 and 5 alternative splice sites have been identified in the C-terminus of Kv7.2 and Kv3.3, respectively. However, the physiological properties of many of these splice variants remain unexplored.

We have developed mouse Kcnq2 and Kcnc3 minigene reporters to investigate the modulation of alternatively spliced exons in Kv7.2 and Kv3.3. Our ongoing research aims to comprehensively assess the physiological characteristics of these splice variants utilizing techniques such as patch clamp in mammalian cells and two-electrode voltage clamp in frog oocytes.

Preliminary studies using these minigene reporters have revealed that splicing factors such as Ptbp2 and Rbfox1 modulate different splicing events, such as splice site selection and cassette exon inclusion of Kv7.2 and Kv3.3.

By exploring the regulation of Kv7.2 and Kv3.3 splicing by diverse splice factors, our research may provide novel insights into the molecular mechanisms underlying BFNC and SCA-13, potentially discovering new targets for therapeutic intervention.