Abstract
While in most patients the identification of genetic alterations causing dystrophinopathies is a relatively straightforward task, a significant number require genomic and transcriptomic approaches that go beyond a routine diagnostic set-up. In this work, we present a Becker Muscular Dystrophy patient with elevated creatinine kinase levels, progressive muscle weakness, mild intellectual disability and a muscle biopsy showing dystrophic features and irregular dystrophin labelling. Routine molecular techniques (Southern-blot analysis, multiplex PCR, MLPA and genomic DNA sequencing) failed to detect a defect in the DMD gene. Muscle DMD transcript analysis (RT-PCR and cDNA-MLPA) showed the absence of exons 75 to 79, seen to be present at the genomic level. These results prompted the application of low-coverage linked-read whole-genome sequencing (WGS), revealing a possible rearrangement involving DMD intron 74 and a region located upstream of the PRDX4 gene. Breakpoint PCR and Sanger sequencing confirmed the presence of a ~8 Mb genomic inversion. Aberrant DMD transcripts were subsequently identified, some of which contained segments from the region upstream of PRDX4. Besides expanding the mutational spectrum of the disorder, this study reinforces the importance of transcript analysis in the diagnosis of dystrophinopathies and shows how WGS has a legitimate role in clinical laboratory genetics.
Highlights
Duchenne and Becker muscular dystrophies (D/BMD) are allelic disorders caused by pathogenic variants in the DMD gene (Xp21.2-p21.1)
These analyses revealed the presence of several aberrant transcripts, corresponding to the use of cryptic splice-sites located both in the DMD gene as well as in the region upstream of PRDX4 (Figure 2)
A comprehensive analytical approach at the genomic and transcript levels enabled the identification of the underlying genetic defect responsible for the patient’s BMD phenotype
Summary
Duchenne and Becker muscular dystrophies (D/BMD) are allelic disorders caused by pathogenic variants in the DMD gene (Xp21.2-p21.1) This gene, the largest known in the human genome, comprises a total of 79 exons and extremely large introns, in all spanning a genomic segment of around 2500 kb. Single nucleotide variants are often found, whereas more complex genomic rearrangements, such as intra-chromosomal inversions or translocations coincident with the DMD gene, are much rarer mutational events (www.LOVD.nl/DMD). These disease-causing variants lead to a deficiency of dystrophin—a cytoskeletal protein expressed mainly in skeletal muscle and heart, and to a lesser extent in the brain and retina (reviewed in [2]). The deficiency of this protein translates into progressive muscle weakness, delay and eventually regression of motor development milestones, respiratory insufficiency and cardiomyopathy
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