A start codon CMT1X mutation associated with transient encephalomyelitis causes complete loss of Cx32

Abstract

X-linked Charcot-Marie-Tooth disease (CMTX1) results from numerous mutations in the GJB1 gene encoding the gap junction protein connexin32 (Cx32) and is one of the commonest forms of inherited neuropathy. Owing to the expression of Cx32 not only in Schwann cells but also in oligodendrocytes, a subset of CMT1X patients develops central nervous system (CNS) clinical manifestations in addition to peripheral neuropathy. While most GJB1 mutations appear to cause peripheral neuropathy through loss of Cx32 function, the cellular mechanisms underlying the CNS manifestations remain controversial. A novel start codon GJB1 mutation (p.Met1Ile) has been found in a CMT1X patient presenting with recurrent episodes of transient encephalomyelitis without apparent signs of peripheral neuropathy. In order to clarify the functional consequences of this mutation, we examined the cellular expression of two different constructs cloned from genomic DNA including the mutated start codon. None of the cloned constructs resulted in detectable expression of Cx32 by immunocytochemistry or immunoblot, although mRNA was produced at normal levels. Furthermore, co-expression with the other major oligodendrocyte connexin, Cx47, had no negative effect on GJ formation by Cx47. Finally, lysosomal and proteasomal inhibition in cells expressing the start codon mutant constructs failed to recover any detection of Cx32 as a result of impaired protein degradation. Our results indicate that the Cx32 start codon mutation is equivalent to a complete loss of the protein with failure of translation, although transcription is not impaired. Thus, complete loss of Cx32 function is sufficient to produce CNS dysfunction with clinical manifestations.

Loss of Coupling Distinguishes GJB1 Mutations Associated with CNS Manifestations of CMT1X from Those Without CNS Manifestations

“…In other words, disruption of this function could be a necessary condition for florid CNS symptoms. This line of reasoning is supported by the recent report of recurrent florid symptoms in a patient carrying a Met1Ile mutation28, predicted to lead to complete loss of Cx32 protein52. This reasoning also suggests the possibility that complete loss of reflexive coupling in Schwann cells may lead to reduced peripheral nerve conduction velocities in patients with CNS + PNS mutants when compared to those with PNS only mutants, a possibility that could potentially be clinically assessed in the future.…”

X linked Charcot-Marie-Tooth disease and multiple sclerosis: emerging evidence for an association

“…In fact, recent studies have found that under the influence of systemic inflammation transdominant effects of mutant Cx32 on other connexins can disrupt oligodendrocyte gap junctions, although oligodendrocyte dysfunction is further exacerbated by intracellularly retained mutants (eg, Thr55Ile) 40. Nevertheless, other studies have recently observed that even mutations associated with a complete loss of Cx32 can produce CNS dysfunction 41. Although a specific genotype-phenotype correlation between Cx32 mutation location (extracellular, intracellular or transmembrane) and WMLs has not been observed, recent studies suggest that the ability of Cx32 mutants to form gap junction plaques and produce adequate levels of junctional coupling in oligodendrocytes may relate to CNS manifestations 42.…”

The Persistent Müllerian Duct Syndrome: An Update Based Upon a Personal Experience of 157 Cases

“…It is highly unlikely that either could allow translation of a functional protein [Mentrup et al, 2011]. Most start codon mutations result in complete absence of the affected protein [Sargiannidou et al, 2015;Jinda et al, 2016].…”