Expression of the human myotonic dystrophy kinase (DMK) gene in transgenic mice.
|Title:||Expression of the human myotonic dystrophy kinase (DMK) gene in transgenic mice.|
|Authors:||Narang, Monica Ajoo.|
|Abstract:||Myotonic dystrophy (DM) is a phenotypically variable, neuromuscular disorder with a number of multisystemic effects. This autosomal dominant disease is associated with the expansion of a triplet (CTG) repeat, located in the 3$\sp\prime$-untranslated region of the DM kinase (DMK) gene. The magnitude of CTG expansion in successive generations corresponds with increasing severity of DM (genetic anticipation). However, the identification of the mutation associated with DM has done little to resolve the molecular effect of CTG expansion on DMK expression or disease determination. To clarify the effect of triplet expansion, DMK protein levels were assessed by immunoblot analysis in DM patient skeletal muscle samples and normal matched controls using a polyclonal antibody generated in our laboratory. Protein levels were found to reflect a role of over-expression of DMK transcript and protein as a mechanism of disease. As a consequence, transgenic mice over-expressing the human DMK gene ("DMK transgenics") and a DMK minigene construct with 100 CTG repeats ("CTG transgenics") were generated in the presence of matrix attachment region (MAR) sequences. DMK transgenic mice show substantial over-expression of human DMK transcript and protein in brain, skeletal muscle, tongue and eye, tissues typically affected in DM. Cryostat sections of' skeletal muscle from DMK transgenic animals revealed diagnostic hallmarks of DM including sarcoplasmic masses, increased centronucleation and type I fiber atrophy. Additionally, primary myoblasts established from these mice showed reduced fusion potential indicating a delay or defect in myoblast differentiation. Furthermore, similar to other effects documented in DM patients, retinal and behaviour alterations were also observed in DMK transgenics. CTG transgenic mice showed intergenerational trinucleotide repeat instability, the degree of which correlated with increasing DM-like pathology in skeletal muscle with each successive generation. These results indicate that over-expression of DMK in mice confers a pathology similar to that observed in DM suggesting that over-expression at the RNA and/or protein level plays a role in the human disease. Furthermore, trinucleotide instability in mice may allow for reproduction of the effects of genetic anticipation.|
|Collection||Thèses, 1910 - 2010 // Theses, 1910 - 2010|