Myotonic dystrophy phenotype without expansion of (CTG)n repeat: An entity distinct from proximal myotonic myopathy (PROMM)?

Myotonic dystrophy (DM) is associated with an expansion of an unstable (CTG)n repeat in the 3' untranslated region of the DM protein kinase (DMPK) gene on chromosome 19q13.3. We studied six patients from two families who showed no expansions of the repeat, in spite of their clinical diagnosis of DM. These patients had multi-systemic manifestations that were distinguishable from those seen in other myotonic disorders, including proximal myotonic myopathy (PROMM). In one additional family, two symptomatic members showed no expanded (CTG)n repeats, while their affected relatives had the expanded repeats. DM haplotype analysis failed to exclude the DMPK locus as a possible site of mutation in each family; however,DMPK mRNA levels were normal. We conclude that a mutation(s) other than the expanded (CTG)n repeat can cause the DM phenotype. The mutation(s) in these families remain(s) to be mapped and characterized.     

Reduction of the DM-associated homeo domain protein (DMAHP) mRNA in different brain areas of myotonic dystrophy patients.

Myotonic dystrophy (DM) is a multisystemic disease caused by expansion of a CTG trinucleotide repeat in the 3' untranslated region of the DMPK protein kinase gene on chromosome 19q13.3. The mechanism by which this expansion causes disease remains unknown. It has been suggested that CTG expansion not only affects the expression of the DMPK gene, but also alters the nuclear RNA metabolism and expression of neighboring genes. DMAHP, which is expressed in various human tissues, including skeletal muscle, heart and brain, is immediately distal to the 3' end of DMPK gene, in a CpG island which contains the CTG repeat. Here we report a 4- to 5-fold reduction of the expression of the DMAHP gene in different brain areas of DM patients. Our results demonstrate that [CTG]n expansion alters the brain DMAHP mRNA expression supporting a dominant-negative effect at the cellular level of DM [CTG]n mutation. The reduced brain expression of DMAHP could explain cerebral impairment in DM patients.     

Ribonuclear inclusions in skeletal muscle in myotonic dystrophy types 1 and 2

Myotonic dystrophy type 1 (DM1) and type 2 (DM2) are caused by genomic expansions of CTG or CCTG repeats. When transcribed, these mutations give rise to repeat expansion RNAs that form nuclear inclusions and compromise the function of myonuclei. Here, we have used in situ hybridization and immunofluorescence to compare DM1 and DM2 and search for proteins that associate with the RNA nuclear (ribonuclear) inclusions. Although muscle disease is generally more severe in DM1, the ribonuclear inclusions were 8- to 13-fold more intense in DM2, implying greater amounts of repeat expansion RNA. Expression of repeat expansion RNA in myoblasts has been implicated in the pathogenesis of congenital DM1. However, we found that repeat expansion RNA is also expressed in myoblasts in DM2, a disorder that has not been associated with a congenital phenotype. Of 10 putative CUG binding proteins tested for colocalization with mutant RNA, only proteins in the muscleblind family were recruited into ribonuclear inclusions. Previous studies have shown activation of the protein kinase, PKR, by expanded CUG repeats in vitro. However, breeding experiments utilizing PKR knockout mice indicate that this kinase is not required for disease pathogenesis in a transgenic mouse model of DM1. We conclude that ribonuclear inclusions are a key feature of the muscle pathology in DM and that sequestration of muscleblind proteins may have a direct role in the disease process.     

Decreased expression of DMPK: correlation with CTG repeat expansion and fibre type composition in myotonic dystrophy type 1

Abstract Myotonic dystrophy type 1 (DM1) is an autosomal dominant disease caused by a trinucleotide repeatexpansion, cytosine-thymine-guanine (CTG)_n, in the 3′ untranslated region of a gene encoding the myotonic dystrophy protein kinase (DMPK). To correlate CTG expansion and protein expression, we studied muscle specimens from 16 adult DM1 patients using three anti-DMPK antibodies for immunoblotting. We estimated the amount of the full-length DMPK (85 kDa) in muscle biopsies from normal controls and from DM1 patients carrying different (CTG)_n expansions. We found that DMPK concentration was decreased to about 50% in DM patients’ muscles; the protein decrease did not seem correlated with the CTG repeat length. However, the fibre type composition in skeletal muscle seemed somehow to affect DMPK decrease, as the lowest level of the enzyme was found in patients with the lowest content of type 1 fibre.     

Effects of age and gene dose on skeletal muscle sodium channel gating in mice deficient in myotonic dystrophy protein kinase

Myotonic muscular dystrophy (DM) is characterized by abnormal skeletal muscle Na channel gating and reduced levels of myotonic dystrophy protein kinase (DMPK). Electrophysiological measurements show that mice deficient in Dmpk have reduced Na currents in muscle. We now find that the Na channel expression level is normal in mouse muscle partially or completely deficient in Dmpk. Reduced current amplitudes are not changed by age or gene dose, and the reduction is not due to changes in macroscopic or microscopic gating kinetics. The mechanism of abnormal membrane excitability in DM may in part be silencing of muscle Na channels due to Dmpk deficiency.     

Progressive skeletal muscle weakness in transgenic mice expressing CTG expansions is associated with the activation of the ubiquitin–proteasome pathway

Myotonic dystrophy type 1 (DM1) is a neuromuscular disease caused by the expansion of a CTG repeat in the DMPK gene and characterised by progressive skeletal muscle weakness and wasting. To investigate the effects of the CTG expansion on the physiological function of the skeletal muscles, we have used a transgenic mouse model carrying the human DM1 region with 550 expanded CTG repeats. Maximal force is reduced in the skeletal muscles of 10-month-old but not in 3-month-old DM1 mice when compared to age-matched non-transgenic littermates. The progressive weakness observed in the DM1 mice is directly related to the reduced muscle mass and muscle fibre size. A significant increase in trypsin-like proteasome activity and Fbxo32 expression is also measured in the DM1 muscles indicating that an atrophic process mediated by the ubiquitin–proteasome pathway may contribute to the progressive muscle wasting and weakness in the DM1 mice.     

Myotonic dystrophy expanded CUG repeats disturb the expression and phosphorylation of tau in PC12 cells

Mental retardation is a main feature of the congenital form of myotonic dystrophy (DM1), however, the molecular mechanisms underlying the central nervous system symptoms of DM1 are poorly understood. We have established a PC12 cell line-based model expressing the DM1 expanded CUG repeats (CTG90 cells) to analyze the effects of this mutation on neuronal functions. Previously, we have reported that CTG90 cells displayed impaired NGF-induced neuronal differentiation. Because disruption of normal expression of the microtubule associated protein tau and neuronal aggregates of hyperphosphorylated tau have been associated with DM1, this study analyzes the behavior of tau in the CTG90 cells. Several alterations of tau were observed in the PC12 cells that express expanded CUG repeats, including a subtle but reproducible reduction in the expression of the tau mRNA splicing isoform containing exon 10, decreased expression of tau and hyperphosphorylation of both tau and high molecular weight tau as well as abnormal nuclear localization of tau phosphorylated at Ser396/404. Interestingly, phosphorylation regulates negatively the activity of tau as microtubule-associated protein. In addition, impaired activity of the Akt/GSK3beta pathway, which phosphorylates tau, was also identified in the CTG90 cells. Besides tau phosphorylation, the Akt/GSK3beta signaling pathway regulates other key processes of PC12 cells, such as apoptosis and neuronal differentiation. Our results indicate that defective neuronal differentiation exhibited by the PC12 cells expressing expanded CUG repeats could be the result of combinatory effects derived from the altered behavior of tau and the impaired activation of the Akt/GSK3beta signaling pathway.     

Clinical case report atypical myopathy in a young girl with 91 CTG repeats in DM1 locus and a positive DM1 family history

Myotonic dystrophy type 1 (DM1) is an autosomal dominant inheritable disease associated with an expansion of CTG repeats in the 3' UTR of the DMPK gene. The subject is an 11-year-old girl with atypical myopathy. Because the proband's family has a positive DM1 history, a molecular-genetic analysis for DM1 was performed. This study showed that proband had a small DMPK expansion (91 CTG repeats) although the observed myopathy would not normally be associated with DM1. These results show how the phenotypic manifestation of DM1 can have unusual symptoms with a completely unexpected relationship to genotype.     

CTG expansion & haplotype analysis in DM1 gene in healthy Iranian population

Myotonic dystrophy type 1 (DM1) is due to an unstable expansion of CTG repeat in the DMPK gene (19q13.3). The CTG repeat is highly polymorphic (5 to 37) in healthy individuals. According to the hypothesis that expanded (CTG)n alleles originated from larger normal alleles, there may exist a correlation between the prevalence of DM1 and the frequency of large size normal alleles. Strong linkage disequilibrium between different length alleles and the three biallelic markers, Alu, Hinf1 and Taq1, has been reported. OBJECTIVE: To determine the distribution of normal alleles, the frequency of larger normal alleles and analysis of the three biallelic markers, in healthy Iranian controls. MATERIAL AND METHODS: Polymerase chain reaction (PCR) was conducted on two hundred unrelated healthy individuals from different ethnic groups living in Iran to determine the size of the alleles. Markers were analyzed by PCR/RFLP on 174 chromosomes from other control healthy individuals. RESULTS: Our data reveals that 23.7% of alleles had 5 CTG repeats and 7.2% of alleles had > 18 CTG repeats. The analysis of haplotypes revealed that 75% of CTG5 and 80% of CTG > 18 had the (+++) haplotype. CONCLUSION: The frequency of alleles with CTG > 18 in Iran is similar to that of Western Europe and Japan.     

Elongation of (CTG)n repeats in myotonic dystrophy protein kinase gene in tumors associated with myotonic dystrophy patients.

Length of (CTG)n triplet repeats in myotonic dystrophy protein kinase gene (DMPK) was estimated in tumors, normal tissues of the same organs, muscles, and leukocytes from three myotonic dystrophy (DM) patients and a non-DM patient. Using cDNA 25 as a probe, a Southern blot analysis of EcoRI- and BglI-digested DNA from these tissues demonstrated the longest expansion of the repeats in the tumors of DM patients. In all tissues from a non-DM patient, the repeat length was confirmed to be stable by PCR analysis. Our data suggest that expanded (CTG)n repeat in tumor tissues may have increased the instability. This study emphasizes the importance of a long-term prospective study on the incidence of tumors in DM to clarify the pathological interrelation between the two entities.     

Possible de novo CTG repeat expansion in the DMPK gene of a patient with cardiomyopathy

CTG triplet repeats of “normal” length in the myotonic dystrophy protein kinase (DMPK) gene have been previously believed to be stable and new pathological expansion was not believed to occur. Here we report possible de novo CTG repeat expansion in the DMPK gene in a patient with cardiomyopathy, who was not diagnosed as having myotonic dystrophy type 1 (DM1) by conventional genetic tests.     

The myotonic dystrophies

Myotonic dystrophy, or dystrophia myotonica (DM), is the most common inherited muscle disorder in adults. DM is a multisystem disease in which the most disabling feature is muscle wasting that begins in the distal limb and cranial muscles. The genetic basis for DM is an expanded CTG repeat in the DMPK gene on chromosome 19. The size of the expanded repeat, and the severity of the disease, tend to increase in successive generations. The mechanism by which this unusual mutation leads to muscle wasting, myotonia, cataracts, heart block, and neurobehavioral abnormalities has not been clearly defined. Identification of the DM gene has made it easier to delineate other DM-like disorders that are clinically and genetically distinct. The most common of these is proximal myotonic myopathy (PROMM), which is characterized by early involvement of proximal limb muscles. The genetic locus for another DM-like disorder, called DM type 2, was recently mapped to chromosome 3.