Epileptic encephalopathy-related Kv2.1 mutants impair clustering but not neuronal excitability
Abstract
The voltage-gated potassium channel Kv2.1, encoded by the epileptic encephalopathy-associated gene KCNB1, is a primary driver of delayed-rectifier K + currents in neurons. These currents contribute to high-frequency firing by preventing depolarization block due to Na + channel inactivation. Wild-type (WT) channels are localized at the soma, proximal dendrites, and the initial segment of the axon, forming aggregates (clusters) via their C-terminal proximal restriction and clustering (PRC) domain. This study investigated the biophysical and functional consequences of two C-terminal truncation mutations (Y529* and R579*), identified in patients with epileptic encephalopathy, which disrupt this critical clustering domain. The mutations induced a change in Kv2.1 subcellular distribution toward distal dendrites and the axon. Cluster formation was impaired, though not abolished, in neurons expressing mutated subunits together with endogenous WT subunits. Consistent with this clustering deficit, single-molecule imaging revealed altered lateral diffusion of mutated channels. While WT channels remained largely immobile (stabilized), mutated forms exhibited intermittent diffusion punctuated by transient immobilization events. The percentage of stabilized trajectories was lower for mutated channels in the soma, dendrites, and distal axons but not in the proximal portion of the axons. The differential diffusive behavior of WT and mutated channels was reproduced using diffusion-capture simulations that accounted for channels with different numbers of PRC domains (i.e., heterotetramers with mutant subunits) and labile scaffolding interactions. Patch-clamp recordings revealed no significant difference in excitability between WT-and mutant-expressing neurons. However, we observed increased firing in both conditions compared to nontransfected neurons. Altogether, our results suggest that (1) mutated subunits form heterotetramers with endogenous WT subunits, (2) several PRC domains per channel are needed to efficiently immobilize Kv2.1 in large clusters, and (3) increasing Kv2.1 channel expression level, independently of clustering, could drive neuronal hyperexcitability.
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BibTeX
@article{paupiah2026epileptic,
title = {{Epileptic encephalopathy-related Kv2.1 mutants impair clustering but not neuronal excitability}},
author = {Anne-Lise Paupiah and Melvyn Ginisty and Capucine Gendre and Marion Russeau and Imane Mouktine and Sabine Levi and Jean-Christophe Poncer and Marianne Renner},
journal = {Biophysical Journal},
year = {2026},
month = oct,
doi = {10.1016/j.bpj.2026.09.009},
url = {https://hal.science/hal-05782753v1},
note = {CC BY-NC-ND 4.0},
}
RIS
TY - JOUR
TI - Epileptic encephalopathy-related Kv2.1 mutants impair clustering but not neuronal excitability
AU - Anne-Lise Paupiah
AU - Melvyn Ginisty
AU - Capucine Gendre
AU - Marion Russeau
AU - Imane Mouktine
AU - Sabine Levi
AU - Jean-Christophe Poncer
AU - Marianne Renner
PY - 2026
DA - 2026/10/07
JO - Biophysical Journal
DO - 10.1016/j.bpj.2026.09.009
UR - https://hal.science/hal-05782753v1
AB - The voltage-gated potassium channel Kv2.1, encoded by the epileptic encephalopathy-associated gene KCNB1, is a primary driver of delayed-rectifier K + currents in neurons. These currents contribute to high-frequency firing by preventing depolarization block due to Na + channel inactivation. Wild-type (WT) channels are localized at the soma, proximal dendrites, and the initial segment of the axon, forming aggregates (clusters) via their C-terminal proximal restriction and clustering (PRC) domain. This study investigated the biophysical and functional consequences of two C-terminal truncation mutations (Y529* and R579*), identified in patients with epileptic encephalopathy, which disrupt this critical clustering domain. The mutations induced a change in Kv2.1 subcellular distribution toward distal dendrites and the axon. Cluster formation was impaired, though not abolished, in neurons expressing mutated subunits together with endogenous WT subunits. Consistent with this clustering deficit, single-molecule imaging revealed altered lateral diffusion of mutated channels. While WT channels remained largely immobile (stabilized), mutated forms exhibited intermittent diffusion punctuated by transient immobilization events. The percentage of stabilized trajectories was lower for mutated channels in the soma, dendrites, and distal axons but not in the proximal portion of the axons. The differential diffusive behavior of WT and mutated channels was reproduced using diffusion-capture simulations that accounted for channels with different numbers of PRC domains (i.e., heterotetramers with mutant subunits) and labile scaffolding interactions. Patch-clamp recordings revealed no significant difference in excitability between WT-and mutant-expressing neurons. However, we observed increased firing in both conditions compared to nontransfected neurons. Altogether, our results suggest that (1) mutated subunits form heterotetramers with endogenous WT subunits, (2) several PRC domains per channel are needed to efficiently immobilize Kv2.1 in large clusters, and (3) increasing Kv2.1 channel expression level, independently of clustering, could drive neuronal hyperexcitability.
SN - 0006-3495
SN - 1542-0086
ER -