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Pathobiologic Mechanisms of Neurodegeneration in Osteopetrosis Derived From Structural and Functional Analysis of 14
Eleonora Di Zanni1, Eleonora Palagano2,3, Laura Lagostena1
1Consiglio Nazionale delle Ricerche, Istituto di Biofisica (CNR-IBF), Dulbecco Telethon Laboratory, Genoa, Italy.
Abstract:
ClC-7 is a chloride-proton antiporter of the CLC protein family. In complex with its accessory protein Ostm-1, ClC-7 localizes to lysosomes and to the osteoclasts' ruffled border, where it plays a critical role in acidifying the resorption lacuna during bone resorption. Gene inactivation in mice causes severe osteopetrosis, neurodegeneration, and lysosomal storage disease. Mutations in the human CLCN7 gene are associated with diverse forms of osteopetrosis. The functional evaluation of ClC-7 variants might be informative with respect to their pathogenicity, but the cellular localization of the protein hampers this analysis. Here we investigated the functional effects of 13 CLCN7 mutations identified in 13 new patients with severe or mild osteopetrosis and a known ADO2 mutation. We mapped the mutated amino acid residues in the homology model of ClC-7 protein, assessed the lysosomal colocalization of ClC-7 mutants and Ostm1 through confocal microscopy, and performed patch-clamp recordings on plasma-membrane-targeted mutant ClC-7. Finally, we analyzed these results together with the patients' clinical features and suggested a correlation between the lack of ClC-7/Ostm1 in lysosomes and severe neurodegeneration. © 2020 American Society for Bone and Mineral Research (ASBMR).
Insights
Mutations in the CLCN7 gene cause osteopetrosis and neurodegeneration. This study links ClC-7/Ostm-1 lysosomal deficiency to severe neurodegeneration in patients with osteopetrosis.
Area of Science:
- Biochemistry
- Genetics
- Cell Biology
Background:
- Chloride-proton antiporter ClC-7, complexed with Ostm-1, is crucial for lysosomal acidification and bone resorption.
- Mutations in the human CLCN7 gene cause osteopetrosis, a bone disorder.
- Evaluating ClC-7 variants' pathogenicity is challenging due to protein localization.
Purpose of the Study:
- To investigate the functional impact of 13 CLCN7 mutations in patients with osteopetrosis.
- To correlate patient clinical features with ClC-7/Ostm-1 localization and function.
- To understand the link between ClC-7/Ostm-1 deficiency and neurodegeneration.
Main Methods:
- Homology modeling to map mutated residues in ClC-7.
- Confocal microscopy to assess lysosomal colocalization of ClC-7 mutants and Ostm1.
- Patch-clamp recordings on plasma-membrane-targeted mutant ClC-7.
Main Results:
- 13 CLCN7 mutations were analyzed in patients with varying osteopetrosis severity.
- Lysosomal colocalization of ClC-7/Ostm-1 was assessed for mutant variants.
- Functional analysis of plasma-membrane-targeted mutants was performed.
Conclusions:
- A correlation was observed between the absence of ClC-7/Ostm-1 in lysosomes and severe neurodegeneration.
- Functional evaluation of CLCN7 variants, combined with clinical data, aids pathogenicity assessment.
- Understanding ClC-7/Ostm-1 function is critical for osteopetrosis and related neurodegenerative disorders.
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