Expanded CUG Repeats Trigger Aberrant Splicing of ClC-1 Chloride Channel Pre-mRNA and Hyperexcitability of Skeletal Muscle in Myotonic Dystrophy

Mankodi, Ami; Takahashi, Masanori; Jiang, Haowei; Beck, Carol L.; Bowers, William J.; Moxley, Richard T.; Cannon, Stephen C.; Thornton, Charles A.

doi:10.1016/s1097-2765(02)00563-4

Cited by 595 publications

(553 citation statements)

References 43 publications

Supporting

Mentioning

519

Contrasting

Unclassified

Order By: Relevance

“…Furthermore, we performed RT-PCR for Clcn1 and Tnnt3 in our mice and uncovered splicing abnormalities (Fig. 2c) similar to those in transgenic mice overexpressing CUG repeats 15 , in the Mbnl1ΔE3 knockout mouse 16 and in individuals with myotonic dystrophy [14][15][16] . Skeletal muscle histology also showed induction of central nuclei, nuclear clumping and fiber size variation as seen in individuals with DM1 (Fig.…”

mentioning

confidence: 62%

“…The cardiac conduction abnormalities observed in these mice are exactly like those seen in up to 70% of individuals with DM1: namely, heart block and atrioventricular node dysfunction 13 . This is the first transgenic mouse model with cardiac conduction Myotonia in DM1 and DM2 is associated with reduced chloride channel (Clcn1, also known as Clc-1) expression and splicing abnormalities of Clcn1 mRNA 14,15 . We found ClC-1 protein significantly reduced or absent from the muscle membranes in mice in which transgene expression was induced (Fig.…”

mentioning

confidence: 99%

“…Myotonia in DM1 and DM2 is associated with reduced chloride channel (Clcn1, also known as Clc-1) expression and splicing abnormalities of Clcn1 mRNA 14,15 . We found ClC-1 protein significantly reduced or absent from the muscle membranes in mice in which transgene expression was induced (Fig.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Reversible model of RNA toxicity and cardiac conduction defects in myotonic dystrophy

et al. 2006

View full text Add to dashboard Cite

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) 1 , which results in nuclear entrapment of the 'toxic' mutant RNA and interacting RNAbinding proteins (such as MBNL1) in ribonuclear inclusions 2 . 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.Common features of adult-onset DM1 include myotonia, progressive skeletal muscle loss, cardiac conduction defects, smooth muscle dysfunction, cataracts and insulin resistance 2 . The normal number of CTG repeats (n = 5 to ~30) is higher (n = 50 to >3,000) in individuals with DM1 (ref. 1 ). Unlike the wild-type transcript, mutant DMPK mRNA forms nuclear aggregates 3,4 and is thought to trigger dominant effects by aberrant interactions with or altered activity of RNA splicing factors, principally members of the muscleblind-like (MBNL) family (such as MBNL1) and the CUG-BP and ETR3-like factor (CELF) family (such as CUG-BP1), leading to abnormal splicing of specific RNAs such as chloride channel (Clcn1), insulin Correspondence should be addressed to M.S.M. (mahadevan@virginia.edu). 4 These authors contributed equally to this work. AUTHOR CONTRIBUTIONSM.S.M., R.S.Y., Q.Y., C.D.F.-M., T.D.B. and L.H.P. performed experimental work and data analysis. S.B. generated the transgene constructs. M.S.M. was responsible for conceptual design and execution. COMPETING INTERESTS STATEMENTThe authors declare that they have no competing financial interests. One potential therapeutic approach in DM1 is to get rid of the toxic RNA from cells. However, it is unclear if this will alleviate the effects of the disease. We used the tetracycline (Tet) inducible system with the reverse tetracycline transactivator (rtTA) to generate double transgenic mice harboring (i) a Tet-responsive, DMPK promoter 10,11 -driven transgene (named GFP-DMPK 3′ UTR) expressing the DMPK 3′ UTR mRNA as part of a GFP transcript, and (ii) a constitutively expressed rtTA transgene (Fig. 1a) Fig. 1). Notably, RNA blots of skeletal muscle RNA showed two major species due to alternative use of polyadenylation signal...

show abstract

mentioning

confidence: 62%

mentioning

confidence: 99%

See 1 more Smart Citation

Reversible model of RNA toxicity and cardiac conduction defects in myotonic dystrophy

et al. 2006

View full text Add to dashboard Cite

show abstract

“…Experimental evidence demonstrates that an alteration in the splicing of several proteins involved in Ca 2+ homeostasis and in excitation-contraction coupling mechanisms may play a pivotal role [6][7][8][9][10]. Myotonia is characterised by a state of pathologically enhanced muscle excitability, in which involuntary trains of action potentials cause a delay in muscle relaxation after contraction [11]. This phenomenon has been associated to the alternative splicing of chloride channels at the muscle fibre level, which determines an alteration of membrane excitability [11].…”

Section: Introductionmentioning

confidence: 99%

Electromechanical delay components during skeletal muscle contraction and relaxation in patients with myotonic dystrophy type 1

Esposito

Cè

Rampichini

et al. 2016

Neuromuscular Disorders

View full text Add to dashboard Cite

This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Highlights Patients with myotonic dystrophy type 1 (DM1) were investigated isometrically  We calculated the electromechanical delays during muscle contraction and relaxation  Delays were partitioned into electrochemical and mechanical components  Measurements reliability was very high in both patients and controls  Delays were longer in DM1, especially during muscle relaxation AbstractThe electromechanical delay during muscle contraction and relaxation can be partitioned into mainly electrochemical and mainly mechanical components by an EMG, mechanomyographic, and force combined approach. Components duration and measurements reliability were investigated during contraction and relaxation in a group of patients with myotonic dystrophy type 1 (DM1, n=13) and in healthy controls (n=13). EMG, mechanomyogram, and force were recorded in DM1 and in age-and body-matched controls from tibialis anterior (distal muscle) and vastus lateralis (proximal muscle) muscles during maximum voluntary and electrically-evoked isometric contractions. The electrochemical and mechanical components of the electromechanical delay during muscle contraction and relaxation were calculated off-line. Maximum strength was significantly lower in DM1 than in controls under both experimental conditions. All electrochemical and mechanical components were significantly longer in DM1 in both muscles.Measurements reliability was very high in both DM1 and controls. The high reliability of the measurements and the differences between DM1 patients and controls suggest that the EMG, mechanomyographic, and force combined approach could be utilized as a valid tool to assess the level of neuromuscular dysfunction in this pathology, and to follow the efficacy of pharmacological or non-pharmacological interventions.

show abstract

“…A multi-step model for DM1 pathogenesis has been proposed [reviewed in reference 2]: (1) the mutant gene is transcribed, giving rise to transcripts that contain an expanded CUG repeat (CUG exp ) [3]; (2) the CUG exp transcripts accumulate in RNA nuclear (ribonuclear) foci [4]; (3) RNA binding proteins, including muscleblind 1 (MBNL1), are sequestered in the ribonuclear foci [5,6]; (4) altered activity of splicing factors, such as MBNL1 and CUG binding protein 1, leads to abnormal alternative splicing for a sub-group of pre-mRNAs [7,8]; and (5) expression of inappropriate splice products leads to symptoms of DM1. For example, CUG exp RNA triggers abnormal alternative splicing of the ClC-1 chloride channel, and the predominant ClC-1 splice products expressed in DM1 muscle are devoid of ion channel activity [9][10][11]. Deficiency of ClC-1 channels contributes to myotonia in DM1, and can be reversed in a transgenic mouse model by overexpressing MBNL1 to levels that exceed the capacity of CUG exp RNA to sequester proteins [12].…”

Section: Introductionmentioning

confidence: 99%

Ribonuclear foci at the neuromuscular junction in myotonic dystrophy type 1

Wheeler

Krym

Thornton

2007

Neuromuscular Disorders

View full text Add to dashboard Cite

In myotonic dystrophy type 1 (DM1) the muscle fibers express RNA containing an expanded CUG repeat (CUG exp ). The CUG exp RNA is retained in the nucleus, forming ribonuclear foci. Splicing factors in the muscleblind (MBNL) family are sequestered in ribonuclear foci, resulting in abnormal regulation of alternative splicing. In extrajunctional nuclei, these effects on splicing regulation lead to reduced chloride conductance and altered insulin receptor signaling. Here we show that CUG exp RNA is also expressed in subsynaptic nuclei of muscle fibers and in motor neurons in DM1, causing sequestration of MBNL1 protein in both locations. In a transgenic mouse model, expression of CUG exp RNA at high levels in extrajunctional nuclei replicates many features of DM1, but the toxic RNA is poorly expressed in subsynaptic nuclei and the mice fail to develop denervation-like features of DM1 myopathology. Our findings indicate that subsynaptic nuclei and motor neurons are at risk for DM1-induced spliceopathy, which may affect function or stability of the neuromuscular junction.

show abstract

Expanded CUG Repeats Trigger Aberrant Splicing of ClC-1 Chloride Channel Pre-mRNA and Hyperexcitability of Skeletal Muscle in Myotonic Dystrophy

Cited by 595 publications

References 43 publications

Reversible model of RNA toxicity and cardiac conduction defects in myotonic dystrophy

Reversible model of RNA toxicity and cardiac conduction defects in myotonic dystrophy

Electromechanical delay components during skeletal muscle contraction and relaxation in patients with myotonic dystrophy type 1

Ribonuclear foci at the neuromuscular junction in myotonic dystrophy type 1

Contact Info

Product

Resources

About