A Recurrent Gain-of-Function Mutation in CLCN6, Encoding the ClC-6 Cl/H+-Exchanger, Causes Early-Onset Neurodegeneration

Maya M. Polovitskaya, Carlo Barbini, Diego Martinelli, Frederike L. Harms, F. Sessions Cole, Paolo Calligari, Gianfranco Bocchinfuso, Lorenzo Stella, Andrea Ciolfi, Marcello Niceta, Teresa Rizza, Marwan Shinawi, Kathleen Sisco, Jessika Johannsen, Jonas Denecke, Rosalba Carrozzo, Daniel J. Wegner, Kerstin Kutsche, Marco Tartaglia, Thomas J. Jentsch

Research output: Contribution to journalArticlepeer-review

17 Scopus citations


Dysfunction of the endolysosomal system is often associated with neurodegenerative disease because postmitotic neurons are particularly reliant on the elimination of intracellular aggregates. Adequate function of endosomes and lysosomes requires finely tuned luminal ion homeostasis and transmembrane ion fluxes. Endolysosomal CLC Cl/H+ exchangers function as electric shunts for proton pumping and in luminal Cl accumulation. We now report three unrelated children with severe neurodegenerative disease, who carry the same de novo c.1658A>G (p.Tyr553Cys) mutation in CLCN6, encoding the late endosomal Cl/H+-exchanger ClC-6. Whereas Clcn6−/− mice have only mild neuronal lysosomal storage abnormalities, the affected individuals displayed severe developmental delay with pronounced generalized hypotonia, respiratory insufficiency, and variable neurodegeneration and diffusion restriction in cerebral peduncles, midbrain, and/or brainstem in MRI scans. The p.Tyr553Cys amino acid substitution strongly slowed ClC-6 gating and increased current amplitudes, particularly at the acidic pH of late endosomes. Transfection of ClC-6Tyr553Cys, but not ClC-6WT, generated giant LAMP1-positive vacuoles that were poorly acidified. Their generation strictly required ClC-6 ion transport, as shown by transport-deficient double mutants, and depended on Cl/H+ exchange, as revealed by combination with the uncoupling p.Glu200Ala substitution. Transfection of either ClC-6Tyr553Cys/Glu200Ala or ClC-6Glu200Ala generated slightly enlarged vesicles, suggesting that p.Glu200Ala, previously associated with infantile spasms and microcephaly, is also pathogenic. Bafilomycin treatment abrogated vacuole generation, indicating that H+-driven Cl accumulation osmotically drives vesicle enlargement. Our work establishes mutations in CLCN6 associated with neurological diseases, whose spectrum of clinical features depends on the differential impact of the allele on ClC-6 function.

Original languageEnglish
Pages (from-to)1062-1077
Number of pages16
JournalAmerican journal of human genetics
Issue number6
StatePublished - Dec 3 2020


  • anion/proton antiport
  • channelopathy
  • chloride channel
  • chloride/proton exchange
  • copper metabolism
  • gain of function
  • gating glutamate
  • luminal pH
  • neurogenic bladder
  • vacuole fusion


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