Direct Evidence that EXP/muscleblind Interacts with CCUG Tetranucleotide Repeats

Myotonic Dystrophy (DM), the most common form of adult-onset muscular dystrophy, comprises at least 2 sub-types, DM1 and DM2. DM1 is caused by the expansion of a CTG repeat located in the 3’ untranslated region (3’UTR) of the DM protein kinase (DMPK) gene. Recently, the expansion of a CCTG tetranucleotide repeat located in the intron of the ZNF9 gene was identified as the mutation responsible for DM2. Since both DM1 and DM2 are caused by the expansion of repetitive sequences, some common factors that interact with these sequences might be involved in the pathogenesis of DM. EXP/muscleblind is a candidate for such factors and is thought to be sequestered by the expanded forms of DM transcripts. Here we performed yeast three-hybrid assays that can detect protein-RNA interactions in vivo to confirm the binding feature of EXP to several kinds of repetitive RNA sequences. We found that both EXP40 and EXP42, splice variants of EXP, showed significant interactions with both CCUG and CUG repeats, but not with UG repeats. These results strongly suggest that EXP might be involved in the pathogenesis of both DM1 and DM2. We found the difference in the intracellular localization of EXP40 and EXP42, though their amino acid sequences differ slightly.