Cellular mechanisms of connexin32 mutations associated with CNS manifestations.
Both oligodendrocytes and myelinating Schwann cells express the gap junction protein connexin32 (Cx32). Mutations in the gene encoding Cx32 (GJB1) cause the X-linked form of Charcot-Marie-Tooth disease (CMTX). Although most CMTX patients do not have clinical central nervous system (CNS) manifestations, subclinical evidence of CNS dysfunction is common. We investigated the cellular effects of a subgroup of GJB1/Cx32 mutations that have been reported to cause clinical CNS dysfunction. We hypothesized that these mutants have dominant-negative effects on other connexins expressed by oligodendrocytes, specifically Cx45. We expressed these and other Cx32 mutants in communication-incompetent as well as Cx45-expressing HeLa cells, and analyzed the transfected cells by immunocytochemistry and immunoblotting. In communication-incompetent cells, the mutants associated with CNS phenotypes failed to reach the cell membrane and were instead retained in the endoplasmic reticulum (A39V, T55I) or Golgi apparatus (M93V, R164Q, R183H), although rare gap junction plaques were found in cells expressing M93V or R183H. In HeLa cells stably expressing Cx45, these Cx32 mutants showed a similar expression pattern, and did not alter the pattern of Cx45 expression. These results indicate that Cx32 mutants that are associated with a CNS phenotype do not interact with Cx45, but may instead have other toxic effects in oligodendrocytes.
Kleopa
Kleopas A
KA
Department of Neurology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA. kleopa@mail.med.upenn.edu
Yum
Sabrina W
SW
Scherer
Steven S
SS
D002450:Cell Communication / Q000235:genetics; D002607:Charcot-Marie-Tooth Disease / Q000503:physiopathology*; D017630:Connexins / Q000235:genetics* / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D017629:Gap Junctions / Q000378:metabolism; D015870:Gene Expression / Q000502:physiology; D006056:Golgi Apparatus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D009097:Multienzyme Complexes / Q000378:metabolism; D016297:Mutagenesis, Site-Directed / Q000502:physiology; D009836:Oligodendroglia / Q000502:physiology*; D010641:Phenotype; D046988:Proteasome Endopeptidase Complex; D021381:Protein Transport / Q000502:physiology; D012583:Schwann Cells / Q000502:physiology; D000097002:Gap Junction beta-1 Protein
D017630:Connexins; D003546:Cysteine Endopeptidases; D009097:Multienzyme Complexes; D046988:Proteasome Endopeptidase Complex; D000097002:Gap Junction beta-1 Protein; C082920:connexin 45