Investigation of Myotonic Dystrophy Kinase Isoform Translocation and Membrane Association

Abstract

Myotonic dystrophy is caused by the expansion of a CTG repeat found in the 3'-untranslated region of the myotonic dystrophy kinase. The mechanism of disease and the role of the kinase are currently obscure. Here we begin the investigation of domain structure/function correlations to aid in determining its normal function. Expressed full-length protein and protein truncated before a C-terminal hydrophobic domain were compared. In vitro, signal peptide function and protection of kinase by microsomal membranes were absent; thus, it is not translocated, as previously proposed. However, full-length kinase expressed in insect cells was found in fractions enriched for membranes and decorated mitochondria. The truncated form was found primarily in the cytosol. The kinase was present as two self-associated, disulfide-linked complexes. The majority of full-length kinase was found in the larger of the two complexes, while almost all of the truncated form was found in the smaller. Thus, the C-terminal region confers a higher order of self-association. Furthermore, full-length kinase expressed in COS-1 cells was present as high molecular weight complex, while the truncated form was present as monomer species. These experiments indicate that the myotonic dystrophy kinase is not membrane-integrated, but that it may have a molecular organization which favors peripheral association with membranes.