Analysis of a Charcot-Marie-Tooth Disease Mutation Reveals an Essential Internal Ribosome Entry Site Element in the Connexin-32 Gene

Alice Hudder, Rudolf Werner
2000 Journal of Biological Chemistry  
A mutation located in the 5-untranslated region (5-UTR) of the nerve-specific connexin-32 mRNA, previously found in a family with Charcot-Marie-Tooth disease (CMTX), was analyzed for its effect on the expression of a reporter gene (luciferase) in transgenic mice and in transfected cells. Whereas both mutant and wild-type genes appeared to be transcribed and spliced efficiently, no luciferase was detected from the mutant in either system, suggesting that the mutation affects translation of the
more » ... NA. When the 5-UTR of nervespecific connexin-32 mRNA was inserted between the two genes of a bicistronic vector and transfected into various cell lines, expression of the second gene was significantly increased. Because the mutant did not facilitate translation of the second gene in the bicistronic mRNA system, this result suggested that the CMTX mutation abolished function of an internal ribosome entry site (IRES) in the 5-UTR of the wild-type connexin-32 mRNA. The CMTX phenotype of the mutant 5-UTR further suggested that the wild-type IRES was essential for the translation of the connexin-32 mRNA in nerve cells. In addition, other sequence elements of the connexin-32 IRES were characterized by mutation analysis. A mutation in either of the first two elements investigated showed loss of IRES function, whereas mutation of a third element showed gain of function.
doi:10.1074/jbc.m005199200 pmid:10931843 fatcat:p7uenzhhgncxzowmgsa6n7wb2u