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. 2020 Apr 10;21(7):2635.
doi: 10.3390/ijms21072635.

Interpretation of the Epigenetic Signature of Facioscapulohumeral Muscular Dystrophy in Light of Genotype-Phenotype Studies

Ana Nikolic  1 , Takako I Jones  2 , Monica Govi  1 , Fabiano Mele  3 , Louise Maranda  4 , Francesco Sera  5 , Giulia Ricci  6 , Lucia Ruggiero  7 , Liliana Vercelli  8 , Simona Portaro  9 , Luisa Villa  10 , Chiara Fiorillo  11 , Lorenzo Maggi  12 , Lucio Santoro  7 , Giovanni Antonini  9 , Massimiliano Filosto  13 , Maurizio Moggio  10 , Corrado Angelini  14 , Elena Pegoraro  15 , Angela Berardinelli  16 , Maria Antonetta Maioli  17 , Grazia D'Angelo  18 , Antonino Di Muzio  19 , Gabriele Siciliano  6 , Giuliano Tomelleri  20 , Maurizio D'Esposito  21 , Floriana Della Ragione  21 , Arianna Brancaccio  21 , Rachele Piras  17 , Carmelo Rodolico  22 , Tiziana Mongini  8 , Frederique Magdinier  23 , Valentina Salsi  1 , Peter L Jones  2 , Rossella Tupler  20   24   25   26
Affiliations

Interpretation of the Epigenetic Signature of Facioscapulohumeral Muscular Dystrophy in Light of Genotype-Phenotype Studies

Ana Nikolic et al. Int J Mol Sci. .

Abstract

Facioscapulohumeral muscular dystrophy (FSHD) is characterized by incomplete penetrance and intra-familial clinical variability. The disease has been associated with the genetic and epigenetic features of the D4Z4 repetitive elements at 4q35. Recently, D4Z4 hypomethylation has been proposed as a reliable marker in the FSHD diagnosis. We exploited the Italian Registry for FSHD, in which FSHD families are classified using the Clinical Comprehensive Evaluation Form (CCEF). A total of 122 index cases showing a classical FSHD phenotype (CCEF, category A) and 110 relatives were selected to test with the receiver operating characteristic (ROC) curve, the diagnostic and predictive value of D4Z4 methylation. Moreover, we performed DNA methylation analysis in selected large families with reduced penetrance characterized by the co-presence of subjects carriers of one D4Z4 reduced allele with no signs of disease or presenting the classic FSHD clinical phenotype. We observed a wide variability in the D4Z4 methylation levels among index cases revealing no association with clinical manifestation or disease severity. By extending the analysis to family members, we revealed the low predictive value of D4Z4 methylation in detecting the affected condition. In view of the variability in D4Z4 methylation profiles observed in our large cohort, we conclude that D4Z4 methylation does not mirror the clinical expression of FSHD. We recommend that measurement of this epigenetic mark must be interpreted with caution in clinical practice.

Keywords: D4Z4 reduced allele; DNA methylation; FSHD; genotype–phenotype correlation; molecular diagnosis.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Assessment of DNA methylation level in facioscapulohumeral muscular dystrophy (FSHD) index cases using MRSE1 (A) and MRSE2 (B) approaches: (A) The crude relationship between the two parameters is not significant (b = −0.028, p = 0.236; R2 = 0.012). The strength of the relationship does not improve when adjusting for the age of patient, as we find no significant association between level of D4Z4 methylation and FSHD score (b = −0.035, p = 0.140; R2 = 0.033). (B) The crude relationship is very weak but statistically significant (b = −0.047, p = 0.048; R2 = 0.036). The strength of the relationship improves adjusting for the age of patient (b=−0.050, p = 0.036, R2 = 0.057).
Figure 2
Figure 2
Comparison of D4Z4 DNA methylation level between FSHD index cases carrying 1–10 D4Z4 repeat units (RU) (FSHD1) or more than 10 D4Z4 RU (FSHD2): A box plot of D4Z4 methylation level assessed with MSRE1 (A) and MSRE2 (B) approaches in carriers of 1–10 or >10 D4Z4 units. The circle indicates an outlier value.
Figure 3
Figure 3
Evaluation of D4Z4 methylation levels in subjects belonging to different clinical categories: A box plot of D4Z4 methylation levels assessed with MSRE1 assay (A) and MSRE2 (B) in probands and relatives stratified over clinical categories (A, B, C, and D).
Figure 4
Figure 4
Evaluation of the efficacy of D4Z4 methylation status as a marker for classifying diseased and non-diseased individuals. The receiver operating characteristic (ROC) curve was plotted to analyze the values of methylation at D4Z4 as a discriminator between affected and unaffected individuals.
Figure 5
Figure 5
Analysis of the D4Z4 methylation status in Family C. (A) Pedigree of family C. The genetic profile (A) or the clinical and epigenetic features (B) of each individual are reported. Filled symbols represent affected people. (C) Genomic DNAs from the selected individuals were analyzed using the 4qA or 4qA-L sodium bisulfite sequencing (BSS) assay. The percent DNA methylation for the Q1 is indicated. Red boxes indicate methylated CpGs, blue boxes indicate unmethylated CpGs, and white boxes indicate no CpG at the expected site.
Figure 6
Figure 6
Analysis of the D4Z4 methylation status in family A. (A) Pedigree of family A. The genetic profile (A) or the clinical and epigenetic features (B) of each individual are reported. Filled symbols represent affected people (C) Genomic DNAs from the selected individuals were analyzed using the 4qA or 4qA-L BSS assay. The percentage DNA methylation for the Q1 is indicated. Red boxes indicate methylated CpGs, blue boxes indicate unmethylated CpGs, and white boxes indicate no CpG at the expected site.
Figure 7
Figure 7
Analysis of the D4Z4 methylation status B. (A) Pedigree of family B. The genetic profile (A) or the clinical and epigenetic features (B) of each individual are reported. Filled symbols represent affected people (C) Genomic DNAs from the selected individuals were analyzed using the 4qA BSS assay. The percentage of DNA methylation for the Q1 is indicated. Red boxes indicate methylated CpGs, blue boxes indicate unmethylated CpGs, and white boxes indicate no CpG at the expected site.
Figure 8
Figure 8
Patients recruitment and their clinical status.
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References

    1. Padberg G.W.A.M. Facioscapulohumeral Disease. Leiden University; Leiden, The Netherlands: 1982.
    1. Mostacciuolo M.L., Pastorello E., Vazza G., Miorin M., Angelini C., Tomelleri G., Galluzzi G., Trevisan C.P. Facioscapulohumeral muscular dystrophy: Epidemiological and molecular study in a north-east Italian population sample. Clin. Genet. 2009;75:550–555. doi: 10.1111/j.1399-0004.2009.01158.x. - DOI - PubMed
    1. Goto K., Nishino I., Hayashi Y.K. Very low penetrance in 85 Japanese families with facioscapulohumeral muscular dystrophy 1A. J. Med. Genet. 2004;41:e12. doi: 10.1136/jmg.2003.008755. - DOI - PMC - PubMed
    1. Nakagawa M., Matsuzaki T., Higuchi I., Fukunaga H., Inui T., Nagamitsu S., Yamada H., Arimura K., Osame M. Facioscapulohumeral Muscular Dystrophy: Clinical Diversity and Genetic Abnormalities in Japanese Patients. Intern. Med. 1997;36:333–339. doi: 10.2169/internalmedicine.36.333. - DOI - PubMed
    1. Jones T.I., Chen J.C.J., Rahimov F., Homma S., Arashiro P., Beermann M.L., King O.D., Miller J.B., Kunkel L.M., Emerson C.P., et al. Facioscapulohumeral muscular dystrophy family studies of DUX4 expression: Evidence for disease modifiers and a quantitative model of pathogenesis. Hum. Mol. Genet. 2012;21:4419–4430. doi: 10.1093/hmg/dds284. - DOI - PMC - PubMed

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