RNA processing is altered in skeletal muscle nuclei of patients affected by myotonic dystrophy

Abstract

Myotonic dystrophies (DMs) are characterised by highly variable clinical manifestations consisting of muscle weakness and atrophy, and a wide spectrum of extramuscular manifestations. In both DM1 and DM2 forms, expanded nucleotide sequences cause the accumulation of mutant transcripts in the nucleus, thus deregulating the function of some RNA-binding proteins and providing a plausible explanation for the multifactorial phenotype of DM patients. However, at the skeletal muscle level, no mechanistic explanation for the muscle wasting has so far been proposed. We therefore performed a study in situ by immunoelectron microscopy on biceps brachii biopsies from DM1, DM2 and healthy subjects, providing the first ultrastructural evidence on the distribution of some nuclear ribonucleoprotein (RNP)-containing structures and molecular factors involved in pre-mRNA transcription and maturation in dystrophic myonuclei. Our results demonstrated an accumulation of splicing and cleavage factors in myonuclei of both DM1 and DM2 patients, suggesting an impairment of post-transcriptional pre-mRNA pathways. The transcription of the expanded sequences in DM myonuclei would therefore hamper functionality of the whole splicing machinery, slowing down the intranuclear molecular trafficking; this would reduce the capability of myonuclei to respond to anabolic stimuli thus contributing to muscle wasting.

Muscleblind-like1 undergoes ectopic relocation in the nuclei of skeletal muscles in myotonic dystrophy and sarcopenia

“…Again, old myonuclei contain significantly higher amount of condensed chromatin than the adults. 37,39 Western blotting results support the immunohistochemical data, showing a significant increase in MBNL1 content both in dystrophic muscles (essentially depending on MBNL1 sequestration in the foci ) and in sarcopenic muscles (where this mainly relates to the protein accumulation in RNP nuclear constituents).…”

“…perichromatin fibrils (PF), perichromatin granules (PG) and interchromatin granules (IG) 36 as well as the molecular factors responsible for pre-mRNA transcription and maturation undergo massive rearrangement in the nuclei of skeletal muscles from DM1 and DM2 patients: 37 the precise intranuclear location of these RNP constituents is an essential prerequisite for pre-mRNA synthesis and processing to correctly take place, and they are considered as highly sensitive markers of nuclear activity 38 . Interestingly, the abnormal RNP distribution in DM muscle cells suggests an impairment of pre-mRNA processing 37 and strongly reminds the nuclear alterations typical of sarcopenia 39-41 i.e ., the loss of muscle mass and function which physiologically occur during ageing 42 and is characterised by myofibre atrophy, fibre size variability and centrally located nuclei 43 . This evidence allowed to hypothesize that common nuclear mechanisms might be responsible for skeletal muscle wasting in sarcopenia and in different muscular pathologies 44-46 …”

Myotonic Dystrophy Type 2: An Update on Clinical Aspects, Genetic and Pathomolecular Mechanism

“…Moreover, in a study in situ by immunoelectron microscopy on muscle biopsies from DM and healthy subjects, an accumulation of splicing and cleavage factors in myonuclei of both DM1 and DM2 patients has been demonstrated suggesting an impairment of post-transcriptional pre-mRNA pathways which could lead to the multiple pathological dysfunctions observed in dystrophic patients [87]. …”

Recent progress in histochemistry and cell biology

“…In skeletal muscle no mechanistic explanation for muscle wasting in DM patients has so far been proposed. Therefore, Malatesta et al (2011) performed in situ studies on biceps brachii biopsies from DM1, DM2 and healthy subjects, applying immunoelectron microscopy. The most accepted hypothesis for both DM1 and DM2 diseases is an accumulation of pathological expanded RNAs [DM1 is caused by an expanded (CTG) n nucleotide sequence in the 3 0 untranslated region of the Dystrophia Myotonic Protein Kinase gene, whereas in DM2 expansion of the tetranucleotidic repeat (CCTG) n in the first intron of the Zinc Finger Protein 9 gene has been detected] in the nucleus, which in turn deregulate the function of RNA-binding proteins and alter mRNA pathways (see recent review in Schoser and Timchenko 2010).…”