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Kv7.3 Compound Heterozygous Variants in Early Onset Encephalopathy Reveal Additive Contribution of C-Terminal Residues to PIP2-Dependent K+ Channel Gating

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Abstract

Over one hundred mutations in the Kv7.2 (KCNQ2) gene encoding for phosphatidylinositol 4,5-bisphosphate (PIP2)-sensitive voltage-gated K+ channel subunits have been identified in early-onset epilepsies with wide phenotypic variability. By contrast, only few mutations in the closely related Kv7.3 (KCNQ3) gene have been reported, mostly associated with typical benign familial neonatal seizures (BFNS). We herein describe a patient affected by early onset epileptic encephalopathy (EOEE) carrying two Kv7.3 missense mutations (p.Val359Leu/V359L and p.Asp542Asn/D542N) in compound heterozygosis, each inherited from an asymptomatic parent. Patch-clamp recordings from transiently transfected CHO cells showed that, when incorporated in physiologically relevant Kv7.2 + Kv7.3 heteromeric channels, expression of Kv7.3 V359L or Kv7.3 D542N subunits failed to affect current density, whereas a significant decrease was instead observed when these mutant subunits were both simultaneously present. Modeling and functional experiments revealed that each variant decreased PIP2-dependent current regulation, with additive effects when the two were co-expressed. Moreover, expression of Kv7.2 subunits carrying the D535N variant previously described in three sporadic EOEE cases prompted functional changes more dramatic when compared to those of the corresponding D542N variant in Kv7.3, but similar to those observed when both Kv7.3 V359L and Kv7.3 D542N subunits were expressed together. Finally, the Kv7 activator retigabine restored channel dysfunction induced by each Kv7.2 or Kv7.3 variant(s). These results provide a plausible molecular explanation for the apparent recessive inheritance of the phenotype in the family investigated, and a rational basis for personalized therapy with Kv7 channel activators in EOEE patients carrying loss-of-function mutations in Kv7.2 or Kv7.3.

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Acknowledgments

The authors are deeply grateful to Dr. Thomas J. Jentsch, Leibniz-Institut für Molekulare Pharmakologie (FMP), Berlin (Germany) for Kv7.2 and Kv7.3 cDNAs; Dr. Alvaro Villarroel, UPV-CSIC, Leioa (ES) for PIP(4)5K lγ cDNA; Dr. Yasushi Okamura, Osaka University, Suita, Osaka (JP) for Dr-VSP-IRES-GFP cDNA.

Funding

The present work was supported by the Telethon Foundation (grant number GGP15113) to MT, the Italian Ministry for University and Research (Project Scientific Independence of Researchers 2014 RBSI1444EM) and the University of Naples “Federico II” and Compagnia di San Paolo in the frame of Program STAR “Sostegno Territoriale alle Attività di Ricerca” (project number 6-CSP-UNINA-120) to FM, and the Italian Ministry of Health Ricerca Finalizzata Giovani Ricercatori 2010 (project GR-2010-2304834 to JCD) and Ricerca Finalizzata Giovani Ricercatori 2016 (project GR-2016-02363337 to JCD and MVS).

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  1. Paolo Ambrosino and Elena Freri contributed equally to this work.

Authors and Affiliations

  1. Department of Medicine and Health Science, University of Molise, Campobasso, Italy

    Paolo Ambrosino, Maria Virginia Soldovieri, Ilaria Mosca, Laura Manocchio & Maurizio Taglialatela

  2. Department of Pediatric Neuroscience, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy

    Elena Freri, Barbara Salis, Francesca Ragona & Tiziana Granata

  3. Unit of Genetics of Neurodegenerative and Metabolic Diseases, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy

    Barbara Castellotti & Cinzia Gellera

  4. Clinical Neurophysiology and Epilepsy Center, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy

    Laura Canafoglia, Silvana Franceschetti & Jacopo C. DiFrancesco

  5. Department of Pharmacy, University of Salerno, Fisciano, Salerno, Italy

    Nunzio Iraci

  6. Department of Neuroscience, University of Naples “Federico II”, Via Pansini 5, 80131, Naples, Italy

    Francesco Miceli & Maurizio Taglialatela

  7. Department of Neurology, San Gerardo Hospital, School of Medicine and Surgery, Milan Center for Neuroscience (NeuroMi), University of Milano-Bicocca, Monza, Italy

    Jacopo C. DiFrancesco

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  1. Paolo Ambrosino
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Ambrosino, P., Freri, E., Castellotti, B. et al. Kv7.3 Compound Heterozygous Variants in Early Onset Encephalopathy Reveal Additive Contribution of C-Terminal Residues to PIP2-Dependent K+ Channel Gating. Mol Neurobiol 55, 7009–7024 (2018). https://doi.org/10.1007/s12035-018-0883-5

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