https://www.cell.com/cell/pdf/0092-8674(92)90154-5.pdf

Molecular basis of myotonic dystrophy: Expansion of a trinucleotide (CTG) repeat at the 3′ end of a transcript encoding a protein kinase family member

Myotonic dystrophy (DM) is the most common inherited neuromuscular disease in adults, with a global incidence of 1 in 8000 individuals. DM is an autosomal dominant, multisystemic disorder characterized primarily by myotonia and progressive muscle weakness. Genomic and complementary DNA probes that map to a 10-kilobase Eco RI genomic fragment from human chromosome 19q13.3 have been used to detect a variable length polymorphism in individuals with DM. Increases in the size of the allele in patients with DM are now shown to be due to an increased number of trinucleotide CTG repeats in the 3' untranslated region of a DM candidate gene. An increase in the severity of the disease in successive generations (genetic anticipation) is accompanied by an increase in the number of trinucleotide repeats. Nearly all cases of DM (98 percent or 253 of 258 individuals) displayed expansion of the CTG repeat region. These results suggest that DM is primarily caused by mutations that generate an amplification of a specific CTG repeat.

https://science.sciencemag.org/content/sci/255/5049/1253.full.pdf

Myotonic dystrophy mutation: an unstable CTG repeat in the 3' untranslated region of the gene

Myotonic dystrophy is the commonest adult form of muscular dystrophy, with an estimated incidence of 1 per 7,500, although this is likely to be an underestimate because of the difficulty of detecting minimally affected individuals. It is a multisystem autosomal dominant disorder of unknown biochemical basis. No case of new mutation has been proven. We have isolated a human genomic clone that detects novel restriction fragments specific to individuals with myotonic dystrophy. A two-allele EcoRI polymorphism is seen in normal individuals, but in most affected individuals one of the normal alleles is replaced by a larger fragment, which varies in length both between unrelated affected individuals and within families. The unstable nature of this region may explain the characteristic variation in severity and age at onset of the disease. A second polymorphism at this locus is in almost complete linkage disequilibrium with myotonic dystrophy, strongly supporting our earlier results which indicated that most cases are descended from one original mutation.

Expansion of an unstable DNA region and phenotypic variation in myotonic dystrophy

Myotonic dystrophy (DM) is the most common form of adult muscular dystrophy, with a prevalence of 2-14 per 100,000 individuals The disease is characterized by progressive muscle weakness and sustained muscle contraction, often with a wide range of accompanying symptoms The age at onset and severity of the disease show extreme variation, both within and between families Despite its clinical variability, this dominant condition segregates as a single locus at chromosome 19q133 in every population studied It is flanked by the tightly linked genetic markers ERCC1 proximally and D19S51 distally; these define the DM critical region We report the isolation of an expressed sequence from this region which detects a DNA fragment that is larger in affected individuals than in normal siblings or unaffected controls The size of this fragment varies between affected siblings, and increases in size through generations in parallel with increasing severity of the disease We postulate that this unstable DNA sequence is the molecular feature that underlies DM

Detection of an unstable fragment of DNA specific to individuals with myotonic dystrophy.

We have analyzed the intracellular localization of transcripts from the myotonin protein kinase (Mt-PK) gene in fibroblasts and muscle biopsies from myotonic dystrophy patients and normal controls. In affected individuals, a trinucleotide expansion in the gene results in the phenotype, the severity of which is proportional to the repeat length. A fluorochrome-conjugated probe (10 repeats of CAG) hybridized specifically to this expanded repeat. Mt-PK transcripts containing CTG repeat expansions were detected in the nucleus as bright foci in DM patient fibroblasts and muscle biopsies, but not from normal individuals. These foci represented transcripts from the Mt-PK gene since they simultaneously hybridized to fluorochrome-conjugated probes to the 5'-end of the Mt-PK mRNA. A single oligonucleotide probe to the repeat and the sense strand each conjugated to different fluorochromes revealed the gene and the transcripts simultaneously, and indicated that these focal concentrations (up to 13 per nucleus) represented predominately posttranscriptional RNA since only a single focus contained both the DNA and the RNA. This concentration of nuclear transcripts was diagnostic of the affected state, and may represent aberrant processing of the RNA.

/pdf/foci-of-trinucleotide-repeat-transcripts-in-nuclei-of-1mnop39z8s.pdf

Foci of trinucleotide repeat transcripts in nuclei of myotonic dystrophy cells and tissues.

In 1918 Fleischer reported that after transmission from one generation to the next, myotonic dystrophy has an earlier onset and is more severe. The hypothesis put forward by Penrose in 1948 that 'anticipation' is caused by bias of index case selection was based on theoretical arguments only and has not been supported by clinical observations. This hypothesis was tested in a clinical and genetic study of 14 families with myotonic dystrophy. Excluding index patients, an earlier onset in the child was found in 98% of 61 parent-child pairs. A greater mean difference in age of onset was found with transmission via the father than via the mother. The comparison of the severity of the disease between parents and children was difficult because of the variation in symptoms but, in general, the disease was more severe in the child than in the parent. The penetrance of the abnormal gene was nearly complete in the 14 families combined, leaving little room for the observation of 'complementary' parent-child pairs in the future. Also in retrospect no complementary parent-child pairs were found in the first generations. Fertility was severely reduced only in the very early onset patient group and this selective infertility cannot be responsible for the total amount of anticipation observed. It is therefore concluded that anticipation may be inherent in the transmission of myotonic dystrophy. This has important consequences for genetic counselling.

H. F. M. Busch

Papers

Anticipation in myotonic dystrophy: fact or fiction?