Two epilepsy-associated variants in KCNA2 (K_V1.2) at position H310 oppositely affect channel functional expression

Two KCNA2 variants (p.H310Y and p.H310R) were discovered in paediatric patients with epilepsy and developmental delay. KCNA2 encodes K(V)1.2-channel subunits, which regulate neuronal excitability. Both gain and loss of K(V)1.2 function cause epilepsy, precluding the prediction of variant effects; and while H310 is conserved throughout the K-V-channel superfamily, it is largely understudied. We investigated both variants in heterologously expressed, human K(V)1.2 channels by immunocytochemistry, electrophysiology and voltage-clamp fluorometry. Despite affecting the same channel, at the same position, and being associated with severe neurological disease, the two variants had diametrically opposite effects on K(V)1.2 functional expression. The p.H310Y variant produced 'dual gain of function', increasing both cell-surface trafficking and activity, delaying channel closure. We found that the latter is due to the formation of a hydrogen bond that stabilizes the active state of the voltage-sensor domain. Additionally, H310Y abolished 'ball and chain' inactivation of K(V)1.2 by K-V beta 1 subunits, enhancing gain of function. In contrast, p.H310R caused 'dual loss of function', diminishing surface levels by multiple impediments to trafficking and inhibiting voltage-dependent channel opening. We discuss the implications for K-V-channel biogenesis and function, an emergent hotspot for disease-associated variants, and mechanisms of epileptogenesis.