High-resolution banding study of an X/4 translocation in a female with Duchenne muscular dystrophy

Summary A rare female case of Duchenne muscular dystrophy with an X/4 translocation was found. Detailed cytogenetic analyses by R-banding and high-resolution G-banding techniques revcaled that the exchange point involved in the translcoation was at the p21.1 band on the X chromosome.     

Turner's syndrome and Duchenne muscular dystrophy in a girl with an X; autosome translocation

A balanced de novo (X;9) translocation was observed in a patient with progressive muscular dystrophy of Duchenne's type (DMD), Turner's syndrome, epilepsy and mental retardation. The involvement of the paternal X is suggested. The assignment of the gene locus for DMD is confirmed on Xp21.     

Uniparental disomy of the entire X chromosome in a female with Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a severe, progressive, X-linked muscle-wasting disorder with an incidence of approximately 1/3,500 male births. Females are also affected, in rare instances. The manifestation of mild to severe symptoms in female carriers of dystrophin mutations is often the result of the preferential inactivation of the X chromosome carrying the normal dystrophin gene. The severity of the symptoms is dependent on the proportion of cells that have inactivated the normal X chromosome. A skewed pattern of X inactivation is also responsible for the clinical manifestation of DMD in females carrying X;autosome translocations, which disrupt the dystrophin gene. DMD may also be observed in females with Turner syndrome (45,X), if the remaining X chromosome carries a DMD mutation. We report here the case of a karyotypically normal female affected with DMD as a result of homozygosity for a deletion of exon 50 of the dystrophin gene. PCR analysis of microsatellite markers spanning the length of the X chromosome demonstrated that homozygosity for the dystrophin gene mutation was caused by maternal isodisomy for the entire X chromosome. This finding demonstrates that uniparental isodisomy of the X chromosome is an additional mechanism for the expression of X-linked recessive disorders. The proband's clinical presentation is consistent with the absence of imprinted genes (i.e., genes that are selectively expressed based on the parent of origin) on the X chromosome.     

Skewed X inactivation in a female MZ twin results in Duchenne muscular dystrophy.

One of female MZ twins presented with muscular dystrophy. Physical examination, creatine phosphokinase levels, and muscle biopsy were consistent with Duchenne muscular dystrophy (DMD). However, because of her sex she was diagnosed as having limb-girdle muscular dystrophy. With cDNA probes to the DMD gene, a gene deletion was detected in the twins and their mother. The de novo mutation which arose in the mother was shown by novel junction fragments generated by HindIII, PstI, or TaqI when probed with cDNA8. Additional evidence of a large gene deletion was given by novel SfiI junction fragments detected by probes p20, J-Bir, and J-66 on pulsed-field gel electrophoresis (PFGE). Immunoblot analysis of muscle from the affected twin showed dystrophin of normal size but of reduced amount. Immunofluorescent visualization of dystrophin revealed foci of dystrophin-positive fibers adjacent to foci of dystrophin-negative fibers. These data indicate that the affected twin is a manifesting carrier of an abnormal DMD gene, her myopathy being a direct result of underexpression of dystrophin. Cytogenetic analysis revealed normal karyotypes, eliminating the possibility of a translocation affecting DMD gene function. Both linkage analysis and DNA fingerprint analysis revealed that each twin has two different X chromosomes, eliminating the possibility of uniparental disomy as a mechanism for DMD expression. On the basis of methylation differences of the paternal and maternal X chromosomes in these MZ twins, we propose uneven lyonization (X chromosome inactivation) as the underlying mechanism for disease expression in the affected female.     

Duchenne muscular dystrophy (DMD) in a female with an X/autosomal translocation: Further evidence that the DMD locus is at Xp21

An isolated case of Duchenne muscular dystrophy in a female who has a de novo t(X;5)(p21;q35) translocation is described. The similarities between this patient and four previously reported females with Duchenne muscular dystrophy are discussed. It is concluded that the locus for Duchenne muscular dystrophy is at Xp21 and, furthermore, that this site may be particularly susceptible both to chromosome breakage and exchange and to gene mutation.     

Human X chromosome markers and Duchenne muscular dystrophy.

Two DNA markers, a random DNA fragment 754 and the cDNA sequence encoding the gene for ornithine transcarbamylase (OTC) have been studied in kindreds segregating for Duchenne muscular dystrophy. 754 and OTC are located close physically to the mutation in the region Xp21 below the breakpoints in two Duchenne females. The genetic distance was found to be approximately 10cM between 754 and DMD (two crossovers in 26 meioses) and to be approximately 10cM between OTC and DMD (two crossovers in 26 meioses). Physical data suggest the order DMD-754-OTC. The frequency of recombination compared to physical distance between these markers and DMD suggests that there may be a hot spot of recombination. The relevance of these observations for the isolation of the DMD mutation and clinical use of these probes is discussed.     

Lowe oculocerebrorenal syndrome in a female with a balanced X;20 translocation: mapping of the X chromosome breakpoint.

A Hispanic girl with Lowe oculocerebrorenal syndrome (OCRL), an X-linked recessive condition characterized by cataracts, glaucoma, mental retardation, and proteinuria, is reported. A balanced X;20 chromosomal translocation with the X chromosome breakpoint at q26.1 was found with high-resolution trypsin-Giemsa banding. Somatic cell hybridization was used to separate the X chromosome derivative and the chromosome 20 derivative in order to position, with respect to the translocation breakpoint, several DNA loci that are linked to the Lowe syndrome locus (Xq24-q26). DXS10 and DXS53 were found to be distal to the breakpoint, whereas DXS37 and DXS42 were located proximal to it. These studies suggest that the OCRL locus lies in the region between these probes. The translocation chromosome originated from an unaffected male without a visible translocation, indicating that the most likely cause of OCRL in this patient is the de novo translocation that disrupted the OCRL locus.     

Molecular analysis of recombination in a family with Duchenne muscular dystrophy and a large pericentric X chromosome inversion.

It has been demonstrated in animal studies that, in animals heterozygous for pericentric chromosomal inversions, loop formation is greatly reduced during meiosis. This results in absence of recombination within the inverted segment, with recombination seen only outside the inversion. A recent study in yeast has shown that telomeres, rather than centromeres, lead in chromosome movement just prior to meiosis and may be involved in promoting recombination. We studied by cytogenetic analysis and DNA polymorphisms the nature of meiotic recombination in a three-generation family with a large pericentric X chromosome inversion, inv(X)(p21.1q26), in which Duchenne muscular dystrophy (DMD) was cosegregating with the inversion. On DNA analysis there was no evidence of meiotic recombination between the inverted and normal X chromosomes in the inverted segment. Recombination was seen at the telomeric regions, Xp22 and Xq27-28. No deletion or point mutation was found on analysis of the DMD gene. On the basis of the FISH results, we believe that the X inversion is the mutation responsible for DMD in this family. Our results indicate that (1) pericentric X chromosome inversions result in reduction of recombination between the normal and inverted X chromosomes; (2) meiotic X chromosome pairing in these individuals is likely initiated at the telomeres; and (3) in this family DMD is caused by the pericentric inversion.     

Mapping of X chromosome translocation breakpoints in females with Duchenne muscular dystrophy with respect to exons of the dystrophin gene

There are rare female patients who suffer from Duchenne or Becker muscular dystrophy because they carry an X;autosome translocation with a breakpoint in the dystrophin gene. We have defined the positions of seven of these breakpoints with respect to exon-containing HindIII fragments detected by dystrophin cDNA. One breakpoint lies between exon-containing Hindlll fragments 7 and 8, five breakpoints between exon-containing HindIII fragments 31 to 41, and one lies close to exon-containing-HindIII fragment 50. The distribution of these and of a further seven translocation breakpoints whose positions are known is compared with that reported for deletions and duplications in affected males.     

Mapping of four translocation breakpoints within the Duchenne muscular dystrophy gene

There are over 20 females with Duchenne or Becker muscular dystrophy (DMD or BMD) who have X-autosome translocations that break the X chromosome within band Xp21. Several of these translocations have been mapped with genomic probes to regions throughout the large (approximately 2000 kb) DMD gene. In this report, a cDNA clone from the 5' end of the gene was used to further map the breakpoints in four X-autosome translocations. A t(X;21) translocation in a patient with BMD and a t(X;1) translocation in a patient with DMD were found to break within a large 110-kb intron between exons 7 and 8. Two other DMD translocations, t(X;5) and t(X;11), were found to break between the first and the second exon of the gene within a presumably large intron (greater than 100 kb). These results demonstrate that all four translocations have disrupted the DMD gene and make it possible to clone and sequence the breakpoints. This will in turn determine whether these translocations occur by chance in these large introns or whether there are sequences that predispose to translocations.     

Germline and somatic mosaicism in a female carrier of Duchenne muscular dystrophy

The family of a male with Duchenne muscular dystrophy (DMD) and a deletion within the dystrophin gene has been studied. Polymerase chain reaction analysis of ectopic mRNA from peripheral blood T+B lymphocytes and the use of (CA) _n repeat polymorphisms in and around the deleted region showed the proband's mother to be both a germline mosaic and a somatic mosaic for the deletion seen in her son. The mutation therefore occurred as a mitotic event early in embryogenesis.     

Molecular analysis of X-autosome translocations in females with Duchenne muscular dystrophy.

To further an understanding of the mechanism of constitutional chromosomal rearrangement, the translocation breakpoints of two X-autosome translocations carried by females with Duchenne or Becker muscular dystrophy have been mapped, cloned and sequenced. Breakpoints were mapped to specific introns within the dystrophin gene and intron sequences spanning the two breakpoints were cloned and used as probes to identify DNA fragments containing the translocation junctions. The junction-containing fragments were cloned after amplification by inverse PCR or single-specific-primer PCR. Sequence through the junctions and the autosomal regions spanning the breakpoints identified the mechanism of rearrangement as non-homologous exchange with minor additions or deletions (0-8 nucleotides) at the breakpoints. Paternal origin of these X-autosome translocations, coupled with evidence for non-transmission of X-autosome translocations through male meiosis suggested that the translocations were the result of a post-meiotic rearrangement in spermiogenesis.     

5'-Nucleotidase in skin fibroblasts from patients with Duchenne muscular dystrophy

The 5'-nucleotidase of plasma membranes of cultured skin fibroblasts from patients with Duchenne muscular dystrophy had a reduced affinity for its substrate, 5'-AMP. The Arrhenius plot of the temperature dependence of this enzyme activity was normal. There was no difference between patients and controls in the specific 5'-nucleotidase activity in the whole cell homogenates.     

miRNome profiling in Duchenne muscular dystrophy; identification of asymptomatic and manifesting female carriers

Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disorder that occurs due to inactivating mutations in DMD gene, leading to muscular dystrophy. Prediction of pathological complications of DMD and the identification of female carriers are important research points that aim to reduce disease burden. Herein, we describe a case of a late DMD patient and his immediate female family members, who all carry same DMD mutation and exhibited varied degrees of symptoms. In our study, we sequenced the whole miRNome in leukocytes and plasma of the family members and results were validated using real-time PCR. Our results highlighted the role of miR-409-3p, miR-424-5p, miR-144-3p as microRNAs that show correlation with the extent of severity of muscular weakness and can be used for detection of asymptomatic carriers. Cellular and circulating levels of miR-494-3p had shown significant increase in symptomatic carriers, which may indicate significant roles played by this miRNA in the onset of muscular weakness. Interestingly, circulating levels of miR-206 and miR-410-3p were significantly increased only in the severely symptomatic carrier. In conclusion, our study highlighted several miRNA species, which could be used in predicting the onset of muscle and/or neurological complications in DMD carriers.     

Molecular analysis of Duchenne and Becker muscular dystrophy

Duchenne muscular dystrophy (DMD) is a progressive and lethal neuromuscular disorder caused by a defective gene on the X chromosome. There is no effective treatment and the biochemical defect is yet unknown. Mapping of the DMD locus to band Xp21 in the short arm of the X chromosome has given rise to strategies to clone the gene from its known location in the chromosome. Two cloning strategies have led to the isolation of a gene that is the largest of any yet described. Portions of the gene are deleted in about 8% of affected males, and rare translocations that disrupt the gene cause the disease in females. The isolation of expressed sequences from the DMD locus will undoubtedly lead to isolation of the gene product and ultimately to an understanding of the basic defect. In the meantime, DNA probes from the DMD locus provide a new and accurate approach for carrier identification and prenatal diagnosis of this dreaded disease.     

Dystrophin analysis in clonal myoblasts derived from a Duchenne muscular dystrophy carrier.

Clonal myogenic cell cultures were established from a potential heterozygote for a mutant Duchenne muscular dystrophy (DMD) gene who was also heterozygous for isozymes of the X-linked enzyme glucose-6-phosphate dehydrogenase. Previous tissue culture studies of this muscle donor demonstrated equal proliferative capacity of myoblasts that had lyonized either the paternal or maternal X-chromosome, indicating that mutation of the DMD gene does not affect growth of myoblasts. If this muscle donor were a gonadal mosaic, this conclusion would be incorrect. In the present study, only those myogenic colonies expressing the glucose-6-phosphate dehydrogenase-A isozyme were found to express dystrophin, indicating that this woman was indeed a heterozygote for DMD. By documenting dystrophin deficiency in a specific population of myogenic cells from this woman, we verify our previous conclusion regarding the normal proliferative capacity of DMD myoblasts. Somatic cell testing of dystrophin expression may offer an alternative to established genetic carrier tests for those women in whom deletions of the DMD are not detectable, whose pedigree structure does not permit linkage analysis, or in whom standard phenotypic analyses are ambiguous.