Helen G. Harley

Unstable DNA sequence in myotonic dystrophy.

A variable DNA sequence has been detected in patients with myotonic dystrophy. We set out to determine whether identification of this specific molecular defect would improve clinical management of patients and families with myotonic dystrophy. 127 affected patients who were studied had an expanded DNA fragment not seen in 73 normal controls. The increase in length of the fragment correlated broadly with disease severity, and we noted expansion of the sequence in successive generations of the same family. Progressive expansion of the affected gene provides a molecular explanation for an apparently earlier onset in successive generations (anticipation) in myotonic dystrophy and supports the role of an unstable repeat sequence as the basis of the defect. The specificity of this finding will assist in accurate diagnosis of myotonic dystrophy and genetic counselling of affected families.

Genomic organization and transcriptional units at the myotonic dystrophy locus.

The genomic structure and apparently complete coding sequence of the myotonic dystrophy protein kinase gene have been determined. The gene contains 15 exons distributed over about 13 kb of genomic DNA. It codes for a protein of 624 amino acids with an N-terminal domain highly homologous to cAMP-dependent serine-threonine protein kinases, an intermediate domain with a high alpha-helical content and weak similarity to various filamentous proteins, and a hydrophobic C-terminal segment. Located in close proximity is a second gene, coding for a transcript of about 3 kb, that is homologous to the gene DMR-N9 in the corresponding mouse locus, but has no homologies to other known genes or proteins. Strong expression of the latter gene in brain suggests that it may have a role in the development of mental symptoms in severe cases of the disease.

Myotonic dystrophy: the correlation of (CTG) repeat length in leucocytes with age at onset is significant only for patients with small expansions

Myotonic dystrophy (DM) was the first of a group of diseases to be identified for which the genetic basis is the expansion of a triplet repeat. Myotonic dystrophy also exhibits anticipation, in which the disease worsens through successive generations. These two features have led many groups to analyse whether a significant negative correlation between triplet repeat length and severity of disease exists. However, the recent molecular finding that two distinct subsets of classically affected DM patients exist, those who export expansion derived DMPK RNA and those who do not, led us to question whether combining data from these two sets of patients is statistically valid. We found that although patients with small expansions showed a significant correlation between age at onset and triplet repeat length, those with larger expansions did not. The theoretical triplet repeat size, which separated the two groups, was also deduced.

Regional localisations and linkage relationships of seven RFLPs and myotonic dystrophy on chromosome 19

SummaryWe have studied the genetic linkage relationships of seven DNA polymorphisms on chromosome 19, with each other and with the myotonic dystrophy locus. The DNA sequences were localised to various regions of the chromosome using translocations in somatic cell hybrids. These results provide the basis for a linkage map of most of chromosome 19, and suggest that the myotonic dystrophy locus is close to the centromere.

Minimal expression of myotonic dystrophy: a clinical and molecular analysis.

A clinical and molecular study is reported of 83 patients considered to be minimally affected with myotonic dystrophy (DM). These had been identified in three ways: 60 subjects were identified on clinical grounds and were divided into those with and those without neuromuscular involvement (groups I and II); nine subjects were at high risk of carrying the DM gene but had a normal phenotype (group III); and 14 were parents of definitely affected patients where neither parent showed clinical abnormalities (group IV). PCR analysis of the CTG repeat in the DM gene showed a range of 70 to 230 repeats for the younger at risk patients in group III, while the asymptomatic gene carriers in group IV had 53 to 60 repeats. The sensitivity of diagnosis by EMG was found to be 39%. For ophthalmic signs this was 97.5%. This suggests that assignment on the basis of minimal clinical features carries a significant error. Molecular analysis, in conjunction with established clinical investigations, should prove valuable in the identification and exclusion of minimal myotonic dystrophy.

Application of a closely linked polymorphism of restriction fragment length to counselling and prenatal testing in families with myotonic dystrophy

The close genetic linkage between the loci for apolipoprotein CII (ApoC2) and myotonic dystrophy makes ApoC2 the closest fully validated marker for prediction of myotonic dystrophy. Application to genetic counselling and presymptomatic and prenatal prediction is reported in seven families with myotonic dystrophy, including one case in which the disorder was excluded prenatally. Only one of the families did not have members with ApoC2 genotypes that allowed prediction, but careful clinical study of older family members was found to be an important factor. ApoC2 typing of families with myotonic dystrophy should be of practical help both in prediction for asymptomatic relatives and for prenatal diagnosis in pregnancies of an affected parent.

Specific molecular prenatal diagnosis for the CTG mutation in myotonic dystrophy.

The results of DNA analysis for the specific mutation of myotonic dystrophy are reported in eight pregnancies (two studied retrospectively) in six families. Four results were normal; in the other four, large DNA expansions were found, comparable to the range seen in severely affected children with congenital onset of the disorder. The results agreed with those obtained by linked DNA markers in the six cases where they were available. We conclude that specific molecular prenatal diagnosis of myotonic dystrophy is feasible, and that an abnormal result may also give a guide to possible severity, though this should be interpreted with caution until greater experience is available.

Localization of a human Na+, K+-ATPase α subunit gene to chromosome 19q12åq13.2 and linkage to the myotonic dystrophy locus

The gene coding for a Na+,K+-ATPase alpha subunit (ATP1A3) has been localized to the q12----q13.2 region of human chromosome 19, potentially close to the myotonic dystrophy (DM) gene. In view of previous studies implicating a Na+,K+-ATPase in the pathology of DM, we have examined the possibility that ATP1A3 is a candidate for the DM locus. Although linked, several clear instances of recombination between ATP1A3 and DM rule out the possibility that mutations in ATP1A3 cause the disease. Examination of multiply informative pedigrees indicates the gene order DM-APOC2-ATP1A3.

Linkage relationships of the insulin receptor gene with the complement component 3, LDL receptor, apolipoprotein C2 and myotonic dystrophy loci on chromosome 19.

SummaryMyotonic dystrophy is associated with disturbances in the insulin response, possibly due to an abnormality of the insulin receptor. Both the myotonic dystrophy (DM) and insulin receptor (INSR) genes are on chromosome 19. Using a cloned gene probe for INSR, we have studied its linkage relationships with the DM locus and other chromosome 19 markers. The results show that INSR is not closely linked to DM, but is located very close to C3, in the region 19pter-19p13.2. This implies that the basic genetic defect which causes DM is not directly responsible for the disturbed insulin response in these patients.

Gene mapping and chromosome 19.

Chromosome 19 is currently the most fully mapped of the smaller chromosomes, with about 40 loci assigned to it (HGM8). Major inherited disorders on this chromosome include myotonic dystrophy and familial hypercholesterolaemia. Other loci include five blood groups, a cluster of apolipoprotein genes, and the receptors for insulin and polio virus. A number of cloned genes and random DNA sequences identify polymorphisms which, together with blood group and other protein polymorphisms, have been used to establish a framework for ordering the loci and estimating genetic distances. Hybrid cell lines allow loci to be assigned to one of eight different regions and a detailed genetic map of the chromosome will be possible in the near future.