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Studies in transgenic mice indicate a loss of connexin32 function in X-linked Charcot-Marie-Tooth disease
1Neurogenetics Branch, National Institute of Neurologic Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892-1250, USA.
Abstract:
X-linked Charcot-Marie-Tooth disease (CMTX) is an inherited demyelinating neuropathy caused by mutations in the gene encoding the gap junction protein connexin32 (Cx32). Despite the identification of over 160 different mutations in the Cx32 coding sequence, it is not known whether the mutations cause the disease manifestations through a loss of Cx32 function or through toxic effects on peripheral nerve. We created transgenic mice with a frameshift mutation at codon 175 (175fs), identified in a large CMTX pedigree. Light microscopic examination of the peripheral nerves from adult transgenic animals showed no pathological features. Western blotting did not show transgenic Cx32 protein in any of the 26 lines, although expression of transgenic messenger RNA was detected by reverse-transcriptase polymerase chain reaction and by ribonuclease protection assay. Our findings indicate that the 175fs mutation results in a loss of Cx32 function, without additional toxic effects.
Insights
X-linked Charcot-Marie-Tooth disease (CMTX) results from connexin32 (Cx32) mutations. This study shows the 175fs mutation causes CMTX via loss of Cx32 function, not toxic effects.
Area of Science:
- Neuroscience
- Genetics
- Molecular Biology
Background:
- X-linked Charcot-Marie-Tooth disease (CMTX) is a demyelinating neuropathy.
- Mutations in the connexin32 (Cx32) gene cause CMTX.
- The mechanism of Cx32 mutations (loss-of-function vs. toxic effects) remains unclear.
Purpose of the Study:
- To investigate the functional consequences of a specific Cx32 frameshift mutation (175fs) found in CMTX patients.
- To determine if the 175fs mutation leads to a loss of Cx32 function or causes toxic effects in peripheral nerves.
Main Methods:
- Generation of transgenic mice carrying the 175fs Cx32 mutation.
- Light microscopy of peripheral nerve pathology in transgenic mice.
- Western blotting to detect Cx32 protein expression.
- RT-PCR and ribonuclease protection assay to detect Cx32 mRNA expression.
Main Results:
- Transgenic mice with the 175fs mutation showed no peripheral nerve pathology.
- No detectable Cx32 protein was found in the nerves of transgenic mice.
- Cx32 mRNA expression was detected, indicating successful gene transfer but absence of functional protein.
Conclusions:
- The 175fs mutation in Cx32 causes X-linked Charcot-Marie-Tooth disease through a loss-of-function mechanism.
- This mutation does not appear to exert its pathogenic effects through additional toxic mechanisms on peripheral nerves.