Alternative titles; symbols
HGNC Approved Gene Symbol: CLCN3
Cytogenetic location: 4q33 Genomic coordinates (GRCh38): 4:169,620,578-169,723,673 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 4q33 | Neurodevelopmental disorder with hypotonia and brain abnormalities | 619512 | Autosomal dominant | 3 |
| Neurodevelopmental disorder with seizures and brain abnormalities | 619517 | Autosomal recessive | 3 |
The CLCN3 gene encodes a chloride (Cl-) channel and transporter that resides on intracellular vesicles of the endosomal/lysosomal system and mediates 2Cl-/H+ exchange, which supports luminal acidification (summary by Duncan et al., 2021).
Borsani et al. (1995) described the isolation and characterization of a human gene (CLCN3) and its mouse homolog (Clcn3) sharing significant sequence and structural similarities to members of the voltage-gated chloride channel family: CLCN1 (118425) and CLCN4 (302910). The CLCN3 gene was found to be expressed primarily in tissues derived from neuroectoderm. Within the brain Clcn3 expression was particularly evident in the hippocampus, olfactory cortex, and olfactory bulb. CLCN3 encodes a 760-amino acid protein that differs by only 2 amino acid residues from the protein encoded by Clcn3. CLCN3 protein also shows a high similarity with GEF1, an integral membrane protein of the yeast Saccharomyces cerevisiae known to be involved in respiration and iron-limited cell growth, and with the predicted protein product of a DNA sequence from the mold Septoria nodorum. The high degree of sequence conservation in distantly related species indicates that the gene has retained a fundamental function throughout evolution.
By analysis of genomic DNAs from human/rodent hybrids containing different human chromosomes, Borsani et al. (1995) mapped the CLCN3 gene to human chromosome 4. Mills et al. (1996) mapped the CLCN3 gene to human 4q32 and to mouse chromosome 8. Using FISH, linkage analysis in the CEPH families, and hybridization to a YAC panel, Taine et al. (1998) concluded that the gene is located in band 4q33.
Neurodevelopmental Disorder With Hypotonia And Brain Abnormalities
In 9 unrelated patients with neurodevelopmental disorder with hypotonia and brain abnormalities (NEDHYBA; 619512), Duncan et al. (2021) identified de novo heterozygous missense variants in the CLCN3 gene (see, e.g., I607T, 600580.0001 and T570I, 600580.0002). The mutations, which were found by exome sequencing, were not present in the gnomAD database. The patients were ascertained through international collaboration after the variants had been identified in different clinical or research settings. Electrophysiologic studies in Xenopus oocytes showed that 2 of the missense variants (I607T and T570I) had reduced voltage-dependent current rectification compared to wildtype. I607T also had decreased current amplitude, and both I607T and T570I had reduced or absent large transient currents. Additional in vitro studies demonstrated that I607T and T570I caused increased inward currents at the acidic pH of late endosomes, possibly resulting from a defect in a feedback mechanism or in the gating process. In contrast, 2 other missense variants tested (V772A and A413V) had current amplitudes and rectification similar to wildtype. Duncan et al. (2021) stated that although the biologic consequences of electrophysiologic alterations observed with some variants remained unclear, missense variants likely cause a gain-of-function effect. The patient with the I607T mutation had the most severe phenotype and died at 23 days of age, suggesting a possible genotype/phenotype correlation. Studies of patient cells were not performed.
Neurodevelopmental Disorder With Seizures And Brain Abnormalities
In 2 sibs (family 10) with neurodevelopmental disorder with seizures and brain abnormalities (NEDSBA; 619517), Duncan et al. (2021) identified a homozygous frameshift mutation in the CLCN3 gene (600580.0003). The mutation, which was found by exome sequencing, segregated with the disorder; each unaffected parent was heterozygous for the mutation. It was not present in the gnomAD database. Functional studies of the variant and studies of patient cells were not performed, but it was predicted to result in a loss of function. The authors noted the phenotypic similarities to mice with homozygous loss of Clcn3 (see ANIMAL MODEL).
CLCN3 is thought to mediate swelling-activated plasma membrane currents, but Stobrawa et al. (2001) showed that this broadly expressed chloride channel is present in endosomal compartments and synaptic vesicles of neurons. While swelling-activated currents are unchanged in mice with disrupted Clcn3, acidification of synaptic vesicles is impaired and there is severe postnatal degeneration of the retina and hippocampus. Electrophysiologic analysis of juvenile hippocampal slices revealed no major functional abnormalities despite slightly increased amplitudes of miniature excitatory postsynaptic currents. Mice almost lacking the hippocampus survive and show several behavioral abnormalities but are still able to acquire motor skills. Clcn3 -/- mice were smaller than their littermates at all ages except immediately after birth. Although they showed overall higher mortality, Clcn3 -/- mice survived for more than a year. The nearly complete loss of hippocampal structures in adult knockout mice was not due to an early developmental defect but rather was caused by a selective degeneration starting about 2 weeks after birth. Clcn3 -/- mice were blind as a consequence of severe retinal degeneration that led to a complete loss of photoreceptors by postnatal day 28. Immunohistochemistry revealed that Clcn3 is concentrated in the inner and outer plexiform layers, the regions of synaptic connections. In summary, Stobrawa et al. (2001) showed that CLCN3 is an intracellular chloride channel. It is also present on synaptic vesicles, where it contributes to their acidification. The phenotype of Clcn3 knockout mice strongly suggests that CLCN3 is not the only chloride channel of synaptic vesicles, and Stobrawa et al. (2001) suggested that other CLC channels may have similar roles.
To determine the physiologic role of CLC3, Yoshikawa et al. (2002) generated Clc3-deficient mice, Clcn3 -/-, by targeted gene disruption. Together with developmental retardation and higher mortality, the null mice showed neurologic manifestations such as blindness, motor coordination deficit, and spontaneous hyperlocomotion. In histologic analysis, the null mice showed a pattern of progressive degeneration of the retina, hippocampus, and ileal mucosa, which resembled the phenotype observed in cathepsin D (CTSD; 116840) knockout mice. A defect in cathepsin D is responsible for congenital lysosomal storage disease with profound neurodegeneration in sheep (Tyynela et al., 2000). This defect results in a lysosomal accumulation of ceroid lipofuscin containing the mitochondrial F1F0 ATPase subunit C (603192). In immunohistochemistry and Western blot analysis, Yoshikawa et al. (2002) found that subunit C was heavily accumulated in the lysosomes of null mice. Furthermore, they detected an elevation in the endosomal pH of the null mice. Results indicated that the neurodegeneration observed in the null mice was caused by an abnormality in the machinery which degrades the cellular protein and was associated with the phenotype of neuronal ceroid lipofuscinosis (CLN1; 256730). The elevated endosomal pH may be an important factor in the pathogenesis of CLN1.
In a female infant (P8) with neurodevelopmental disorder with hypotonia and brain abnormalities (NEDHYBA; 619512) who died at 23 days of age, Duncan et al. (2021) identified a de novo heterozygous c.1820T-C transition (c.1820T-C, NM_173872.3) in the CLCN3 gene, resulting in an ile607-to-thr (I607T) substitution. The variant, which was found by exome sequencing, was not present in the gnomAD database. In vitro electrophysiologic studies in Xenopus oocytes transfected with the mutation showed that it caused decreased current amplitude, impaired current rectification, absent transient current, and increased current at a low pH compared to controls. However, similar studies in HEK293 cells transfected with the mutation showed different results (increased amplitude and normal rectification), suggesting that functional properties are dependent on the expression system. Overall, the variant caused electrophysiologic abnormalities consistent with a gain of function. Studies of patient cells were not performed.
In 2 unrelated girls (P6 and P7) with neurodevelopmental disorder with hypotonia and brain abnormalities (NEDHYBA; 619512), Duncan et al. (2021) identified a de novo heterozygous c.1709C-T transition (c.1709C-T, NM_173872.3) in the CLCN3 gene, resulting in a thr570-to-ile (T570I) substitution. The variant, which was found by exome sequencing, was not present in the gnomAD database. In vitro electrophysiologic studies in cells transfected with the mutation showed that it altered certain current properties, most consistent with a gain of function. Studies of patient cells were not performed.
In 2 sibs (family 10) with neurodevelopmental disorder with seizures and brain abnormalities (NEDSBA; 619517), Duncan et al. (2021) identified a homozygous 4-bp deletion (c.336_339del, NM_173872.3) in the CLCN3 gene, resulting in a frameshift and premature termination (Lys112AsnfsTer6). The mutation, which was found by exome sequencing, segregated with the disorder; each unaffected parent was heterozygous for the mutation. It was not present in the gnomAD database. Functional studies of the variant and studies of patient cells were not performed, but it was predicted to result in a loss of function.
Borsani, G., Rugarli, E. I., Taglialatela, M., Wong, C., Ballabio, A. Characterization of a human and murine gene (CLCN3) sharing similarities to voltage-gated chloride channels and to a yeast integral membrane protein. Genomics 27: 131-141, 1995. [PubMed: 7665160] [Full Text: https://doi.org/10.1006/geno.1995.1015]
Duncan, A. R., Polovitskaya, M. M., Gaitan-Penas, H., Bertelli, S., VanNoy, G. E., Grant, P. E., O'Donnell-Luria, A., Valivullah, Z., Lovgren, A. K., England, E. M., Agolini, E., Madden, J. A., and 28 others. Unique variants in CLCN3, encoding an endosomal anion/proton exchanger, underlie a spectrum of neurodevelopmental disorders. Am. J. Hum. Genet. 108: 1450-1465, 2021. [PubMed: 34186028] [Full Text: https://doi.org/10.1016/j.ajhg.2021.06.003]
Mills, K. A., Mathews, K. D., Scherpbier-Heddema, T., Buetow, K. H., Baldini, A., Ballabio, A., Borsani, G. Genetic and physical mapping of a voltage-dependent chloride channel gene to human 4q32 and to mouse 8. Genomics 36: 374-376, 1996. [PubMed: 8812471] [Full Text: https://doi.org/10.1006/geno.1996.0480]
Stobrawa, S. M., Breiderhoff, T., Takamori, S., Engel, D., Schweizer, M., Zdebik, A. A., Bosl, M. R., Ruether, K., Jahn, H., Draguhn, A., Jahn, R., Jentsch, T. J. Disruption of ClC-3, a chloride channel expressed on synaptic vesicles, leads to a loss of the hippocampus. Neuron 29: 185-196, 2001. [PubMed: 11182090] [Full Text: https://doi.org/10.1016/s0896-6273(01)00189-1]
Taine, L., Coupry, I., Boisseau, P., Saura, R., Lacombe, D., Arveiler, B. Refined localisation of the voltage-gated chloride channel, CLCN3, to 4q33. Hum. Genet. 102: 178-181, 1998. [PubMed: 9521585] [Full Text: https://doi.org/10.1007/s004390050673]
Tyynela, J., Sohar, I., Sleat, D. E., Gin, R. M., Donnelly, R. J., Baumann, M., Haltia, M., Lobel, P. A mutation in the ovine cathepsin D gene causes a congenital lysosomal storage disease with profound neurodegeneration. EMBO J. 19: 2786-2792, 2000. [PubMed: 10856224] [Full Text: https://doi.org/10.1093/emboj/19.12.2786]
Yoshikawa, M., Uchida, S., Ezaki, J., Rai, T., Hayama, A., Kobayashi, K., Kida, Y., Noda, M., Koike, M., Uchiyama, Y., Marumo, F., Kominami, E., Sasaki, S. CLC-3 deficiency leads to phenotypes similar to human neuronal ceroid lipofuscinosis. Genes Cells 7: 597-605, 2002. [PubMed: 12059962] [Full Text: https://doi.org/10.1046/j.1365-2443.2002.00539.x]