RESUMEN
Myotonic dystrophy (DM1), the most common muscular dystrophy in adults, is caused by an expanded (CTG)n tract in the 3' UTR of the gene encoding myotonic dystrophy protein kinase (DMPK), which results in nuclear entrapment of the 'toxic' mutant RNA and interacting RNA-binding proteins (such as MBNL1) in ribonuclear inclusions. It is unclear if therapy aimed at eliminating the toxin would be beneficial. To address this, we generated transgenic mice expressing the DMPK 3' UTR as part of an inducible RNA transcript encoding green fluorescent protein (GFP). We were surprised to find that mice overexpressing a normal DMPK 3' UTR mRNA reproduced cardinal features of myotonic dystrophy, including myotonia, cardiac conduction abnormalities, histopathology and RNA splicing defects in the absence of detectable nuclear inclusions. However, we observed increased levels of CUG-binding protein (CUG-BP1) in skeletal muscle, as seen in individuals with DM1. Notably, these effects were reversible in both mature skeletal and cardiac muscles by silencing transgene expression. These results represent the first in vivo proof of principle for a therapeutic strategy for treatment of myotonic dystrophy by ablating or silencing expression of the toxic RNA molecules.
Asunto(s)
Miocardio/metabolismo , Miotonía/fisiopatología , Distrofia Miotónica/genética , Distrofia Miotónica/fisiopatología , ARN/toxicidad , Regiones no Traducidas 3' , Animales , Modelos Animales de Enfermedad , Electrocardiografía , Electromiografía , Silenciador del Gen , Proteínas Fluorescentes Verdes/metabolismo , Inmunohistoquímica , Hibridación Fluorescente in Situ , Ratones , Ratones Transgénicos , Músculo Esquelético/química , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiopatología , Miocardio/química , Distrofia Miotónica/etiología , Distrofia Miotónica/metabolismo , Proteína Quinasa de Distrofia Miotónica , Unión Proteica , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , ARN Mensajero/análisis , Transgenes , Expansión de Repetición de TrinucleótidoRESUMEN
Myotonic muscular dystrophy (DM1) is the most common inherited neuromuscular disorder in adults and is considered the first example of a disease caused by RNA toxicity. Using a reversible transgenic mouse model of RNA toxicity in DM1, we provide evidence that DM1 is associated with induced NKX2-5 expression. Transgene expression resulted in cardiac conduction defects, increased expression of the cardiac-specific transcription factor NKX2-5 and profound disturbances in connexin 40 and connexin 43. Notably, overexpression of the DMPK 3' UTR mRNA in mouse skeletal muscle also induced transcriptional activation of Nkx2-5 and its targets. In human muscles, these changes were specific to DM1 and were not present in other muscular dystrophies. The effects on NKX2-5 and its downstream targets were reversed by silencing toxic RNA expression. Furthermore, using Nkx2-5+/- mice, we show that NKX2-5 is the first genetic modifier of DM1-associated RNA toxicity in the heart.