Allele-specific DNA hypomethylation characterises FSHD1 and FSHD2

Background Facioscapulohumeral muscular dystrophy (FSHD) is associated with an epigenetic defect on 4qter. Two clinically indistinguishable forms of FSHD are known, FSHD1 and FSHD2. FSHD1 is caused by contraction of the highly polymorphic D4Z4 macrosatellite repeat array on chromosome 4q35. FSHD2 is caused by pathogenic mutations of the SMCHD1 gene. Both genetic defects lead to D4Z4 DNA hypomethylation. In the presence of a polymorphic polyadenylation signal (PAS), DNA hypomethylation leads to inappropriate expression of the D4Z4-encoded DUX4 transcription factor in skeletal muscle. Currently, hypomethylation is not diagnostic per se because of the interference of non-pathogenic arrays and the lack of information about the presence of DUX4-PAS. Methods We investigated, by bisulfite sequencing, the DNA methylation levels of the region distal to the D4Z4 array selectively in PAS-positive alleles. Results Comparison of FSHD1, FSHD2 and Control subjects showed a highly significant difference of methylation levels in all CpGs tested. Importantly, using a cohort of 112 samples, one of these CpGs (CpG6) is able to discriminate the affected individuals with a sensitivity of 0.95 supporting this assay potential for FSHD diagnosis. Moreover, our study showed a relationship between PAS-specific methylation and severity of the disease. Conclusions These data point to the CpGs distal to the D4Z4 array as a critical region reflecting multiple factors affecting the epigenetics of FSHD. Additionally, methylation analysis of this region allows the establishment of a rapid and sensitive tool for FSHD diagnosis.

[1]  Wolfram Kress,et al.  Diagnostic approach for FSHD revisited: SMCHD1 mutations cause FSHD2 and act as modifiers of disease severity in FSHD1 , 2014, European Journal of Human Genetics.

[2]  O. King,et al.  Individual epigenetic status of the pathogenic D4Z4 macrosatellite correlates with disease in facioscapulohumeral muscular dystrophy , 2015, Clinical Epigenetics.

[3]  Daniel G. Miller,et al.  Inter-individual differences in CpG methylation at D4Z4 correlate with clinical variability in FSHD1 and FSHD2. , 2015, Human molecular genetics.

[4]  O. King,et al.  Identifying diagnostic DNA methylation profiles for facioscapulohumeral muscular dystrophy in blood and saliva using bisulfite sequencing , 2014, Clinical Epigenetics.

[5]  A. Verbeek,et al.  Population-based incidence and prevalence of facioscapulohumeral dystrophy , 2014, Neurology.

[6]  F. Puppo,et al.  Differential DNA methylation of the D4Z4 repeat in patients with FSHD and asymptomatic carriers , 2014, Neurology.

[7]  M. Kyba,et al.  A focal domain of extreme demethylation within D4Z4 in FSHD2 , 2013, Neurology.

[8]  Daniel G. Miller,et al.  Digenic inheritance of an SMCHD1 mutation and an FSHD-permissive D4Z4 allele causes facioscapulohumeral muscular dystrophy type 2 , 2012, Nature Genetics.

[9]  L. Salviati,et al.  Patients with a phenotype consistent with facioscapulohumeral muscular dystrophy display genetic and epigenetic heterogeneity , 2011, Journal of Medical Genetics.

[10]  J. Statland,et al.  Facioscapulohumeral muscular dystrophy: molecular pathological advances and future directions. , 2011, Current opinion in neurology.

[11]  M. Tarnopolsky,et al.  Clinical features of facioscapulohumeral muscular dystrophy 2 , 2010, Neurology.

[12]  Daniel G. Miller,et al.  A Unifying Genetic Model for Facioscapulohumeral Muscular Dystrophy , 2010, Science.

[13]  P. de Knijff,et al.  Worldwide population analysis of the 4q and 10q subtelomeres identifies only four discrete interchromosomal sequence transfers in human evolution. , 2010, American journal of human genetics.

[14]  R. Frants,et al.  Common epigenetic changes of D4Z4 in contraction‐dependent and contraction‐independent FSHD , 2009, Human mutation.

[15]  Daniel G. Miller,et al.  RNA transcripts, miRNA-sized fragments and proteins produced from D4Z4 units: new candidates for the pathophysiology of facioscapulohumeral dystrophy. , 2009, Human molecular genetics.

[16]  Rune R. Frants,et al.  Specific Loss of Histone H3 Lysine 9 Trimethylation and HP1γ/Cohesin Binding at D4Z4 Repeats Is Associated with Facioscapulohumeral Dystrophy (FSHD) , 2009, PLoS genetics.

[17]  P. de Knijff,et al.  Specific sequence variations within the 4q35 region are associated with facioscapulohumeral muscular dystrophy. , 2007, American journal of human genetics.

[18]  D. Smeets,et al.  Hypomethylation is restricted to the D4Z4 repeat array in phenotypic FSHD , 2007, Neurology.

[19]  E. Ricci,et al.  Variable hypomethylation of D4Z4 in facioscapulohumeral muscular dystrophy , 2005, Annals of neurology.

[20]  G. V. Ommen,et al.  Hypomethylation of D4Z4 in 4q-linked and non-4q-linked facioscapulohumeral muscular dystrophy , 2003, Nature Genetics.

[21]  Long-Cheng Li,et al.  MethPrimer: designing primers for methylation PCRs , 2002, Bioinform..

[22]  E. Bakker,et al.  Complete allele information in the diagnosis of facioscapulohumeral muscular dystrophy by triple DNA analysis , 2001, Annals of neurology.

[23]  G. Deidda,et al.  Direct detection of 4q35 rearrangements implicated in facioscapulohumeral muscular dystrophy (FSHD). , 1996, Journal of medical genetics.

[24]  R. Tupler,et al.  Monosomy of distal 4q does not cause facioscapulohumeral muscular dystrophy. , 1996, Journal of medical genetics.

[25]  J E Hewitt,et al.  FSHD associated DNA rearrangements are due to deletions of integral copies of a 3.2 kb tandemly repeated unit. , 1993, Human molecular genetics.

[26]  C. Wijmenga,et al.  Chromosome 4q DNA rearrangements associated with facioscapulohumeral muscular dystrophy , 1992, Nature genetics.

[27]  P Halonen,et al.  [Facioscapulohumeral muscular dystrophy]. , 1990, Duodecim; laaketieteellinen aikakauskirja.