{
  "abstract": "Due to the complexity of the genomic region, the relationship between genotype, phenotype and epigenetics in facioscapulohumeral muscular dystrophy remains incompletely understood, and diagnostics are limited in precision. In this study, we applied Oxford Nanopore Technology long-read sequencing to characterize FSHD1 and FSHD2 patients with common and previously confirmed genotypes (n = 14). This approach enabled the simultaneous determination of allele-specific methylation profiles, haplotypes and repeat lengths. Our results confirm the presence of an asymmetric methylation gradient increasing from the proximal to the distal end of 4q35 repeat arrays with a steeper slope in FSHD1 (2.5-8.1%/repeat unit) compared to the two FSHD2 patients (0.3 and 1.2%/repeat unit) studied. The slope is, therefore, a potential marker to differentiate both types of facioscapulohumeral muscular dystrophy, being less biased by repeat length compared to parameters determined in current bisulfite sequencing-based methylation assays. By determining distal methylation allele-specifically, long-read sequencing overcomes a major limitation of such assays. Distal methylation of pathogenic (range 22-45%) and non-pathogenic repeat arrays (range 77-87%) (P < 0.0001) is clearly separated and shows a stronger correlation with age-corrected clinical severity (r = -0.81, 95% CI: -0.48 to -0.94) compared to studies based on bisulfite sequencing. Overall, this strengthens distal methylation as a superior diagnostic marker of facioscapulohumeral muscular dystrophy disease status. Detailed clinical and genetic characterization of a second cohort (n = 20) comprising patients with complex genotypes (n = 11) and FSHD2 (n = 9) was performed using our bisulfite sequencing-based methylation profile analysis, complementary methods and in three cases, long-read sequencing. Based on these results, we highlight underrecognized patient groups at risk of misdiagnosis when methylation analysis is not part of the diagnostic workup: (i) patients carrying a 4qA166 repeat array; (ii) patients with underrepresented population backgrounds; (iii) facioscapulohumeral muscular dystrophy patients carrying 11 repeat units in the absence of global hypomethylation; and (iv) FSHD2 patients without a detectable causative variant in known epigenetic suppressor genes (SMCHD1, DNMT3B and LRIF1). We hypothesize that unknown epigenetic modifiers explain the penetrance of repeat arrays with 8-10 repeat units in the European and North American populations, the pathogenicity of some repeat arrays with 11 repeat units and the hypomethylation of the entire 4q35 repeat array in some FSHD2 patients without a detected variant in known epigenetic suppressor genes. From our results, recommendations for standard diagnostics are derived. Finally, we hypothesize a preliminary model linking DNA methylation, known genetic parameters and disease severity.",
  "abstract_shown": true,
  "affiliations": [
    "MGZ - Medizinisch Genetisches Zentrum, Munich 80335, Germany.",
    "Friedrich Baur Institute at the Department of Neurology, LMU University Hospital, Ludwig-Maximilians-Universität München, Munich 80336, Germany.",
    "Department of Medicine IV, LMU University Hospital, Ludwig-Maximilians-Universität München, Munich 80336, Germany.",
    "Department of Neurology, BG University Hospital Bergmannsheil, Ruhr-University Bochum, Bochum 44789, Germany.",
    "BG University Hospital Bergmannsheil, Heimer Institute for Muscle Research, Bochum 44789, Germany.",
    "Department of Neurology with Experimental Neurology, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt Universität zu Berlin, and Berlin Institute of Health (BIH), Berlin 10117, Germany.",
    "Institute of Neurogenomics, Helmholtz Center Munich, Neuherberg 85764, Germany.",
    "Department of Neurology and Mannheim Center for Translational Neuroscience, Medical Faculty Mannheim, Heidelberg University, Mannheim 68167, Germany.",
    "Department of Neurology, University Hospital Carl Gustav Carus at TUD Dresden University of Technology, Dresden 01307, Germany.",
    "Department of Neurology, University Hospital Linz, Linz 4021, Austria.",
    "Institute for Clinical Genetics, University Hospital Carl Gustav Carus at TUD Dresden University of Technology, Dresden 01307, Germany.",
    "Department of Neuropediatrics, University Hospital Carl Gustav Carus at TUDDresden University of Technology, Dresden 01307, Germany.",
    "Department of Neurology and Center for Translational Neuro- and Behavioral Sciences (C-TNBS), University Hospital Essen, Essen 45147, Germany.",
    "Institute of Human Genetics, University Hospital Heidelberg, Heidelberg 69120, Germany.",
    "Neurologie Mingers, Übach-Palenberg 52531, Germany.",
    "Institute for Human Genetics, Hannover Medical School, Hannover 30625, Germany.",
    "Mitteldeutscher Praxisverbund Humangenetik, Leipzig 04103, Germany.",
    "Clinical Genetics and Genomic Medicine, University Hospital Würzburg, Würzburg 97080, Germany."
  ],
  "article_type": "research article",
  "arxiv": null,
  "author_total": 35,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "Florentine Scharf"
    },
    {
      "affs": [
        0,
        1,
        2
      ],
      "name": "Morghan C Lucas",
      "orcid": "0000-0001-7654-9137"
    },
    {
      "affs": [
        0
      ],
      "name": "Stefanie Gehling"
    },
    {
      "affs": [
        3,
        4,
        5
      ],
      "name": "Felix Kleefeld"
    },
    {
      "affs": [
        0
      ],
      "name": "Veronika Schönrock"
    },
    {
      "affs": [
        0,
        6
      ],
      "name": "Anna Benet-Pagès"
    },
    {
      "affs": [
        7
      ],
      "name": "Hans-Werner Rausch"
    },
    {
      "affs": [
        7
      ],
      "name": "Lucas Schirmer",
      "orcid": "0000-0001-7142-4116"
    },
    {
      "affs": [
        8
      ],
      "name": "Jochen Schäfer"
    },
    {
      "affs": [
        9
      ],
      "name": "Annika Saak"
    },
    {
      "affs": [
        10
      ],
      "name": "Joseph Porrmann"
    },
    {
      "affs": [
        10
      ],
      "name": "Sylvia Hütter"
    },
    {
      "affs": [
        11
      ],
      "name": "Christina Emde"
    },
    {
      "affs": [
        11
      ],
      "name": "Anne Boblest"
    },
    {
      "affs": [
        12
      ],
      "name": "Svenja Neuhoff"
    },
    {
      "affs": [
        12
      ],
      "name": "Tim Hagenacker"
    },
    {
      "affs": [
        13
      ],
      "name": "Sebastian Burkart"
    },
    {
      "affs": [
        14
      ],
      "name": "Sebastian Mingers"
    },
    {
      "affs": [
        15
      ],
      "name": "Ann-Cathrine Berking"
    },
    {
      "affs": [
        15
      ],
      "name": "Lisa Olfe"
    },
    {
      "affs": [
        15
      ],
      "name": "Corinna Hendrich"
    },
    {
      "affs": [
        16
      ],
      "name": "Kristin Schröck"
    },
    {
      "affs": [
        17
      ],
      "name": "Elizabeta Sauer"
    },
    {
      "affs": [
        5
      ],
      "name": "Agata A Mossakowski"
    },
    {
      "affs": [
        0
      ],
      "name": "Kai Sendelbach"
    },
    {
      "affs": [
        0
      ],
      "name": "Udo Koehler"
    },
    {
      "affs": [
        0
      ],
      "name": "Kerstin Becker"
    },
    {
      "affs": [
        0
      ],
      "name": "Tobias Bartolomaeus"
    },
    {
      "affs": [
        0,
        2
      ],
      "name": "Ariane Hallermayr",
      "orcid": "0000-0002-6503-1874"
    },
    {
      "affs": [
        0
      ],
      "name": "Barbara Klink"
    },
    {
      "affs": [
        0
      ],
      "name": "Teresa Neuhann"
    },
    {
      "affs": [
        1
      ],
      "name": "Maggie C Walter"
    },
    {
      "affs": [
        1
      ],
      "name": "Benedikt Schoser",
      "orcid": "0000-0002-2757-8131"
    },
    {
      "affs": [
        0,
        1
      ],
      "name": "Angela Abicht"
    },
    {
      "affs": [
        0,
        1
      ],
      "name": "Hannes Erdmann",
      "orcid": "0000-0002-2620-1850"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/dk1sav6rkah04.bib",
  "cited_by": 0,
  "doi": "10.1093/braincomms/fcag370",
  "fulltext_url": "https://europepmc.org/articles/PMC13685627",
  "id": "dk1sav6rkah04",
  "issns": [
    "2632-1297"
  ],
  "landing_url": "https://europepmc.org/article/MED/42909895",
  "language": "eng",
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "openalex",
      "1312|Muscle Physiology and Disorders"
    ]
  ],
  "openalex": "W7221357826",
  "pdf_url": "https://europepmc.org/articles/PMC13685627?pdf=render",
  "pmcid": "PMC13685627",
  "pmid": "42909895",
  "published": "2026-10-10",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/dk1sav6rkah04.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:20:10Z",
      "native_id": "MED:42909895",
      "source": "europepmc",
      "url": "https://europepmc.org/article/MED/42909895"
    }
  ],
  "subjects": [
    {
      "field": "biochemistry",
      "origin": "openalex:1312|Muscle Physiology and Disorders",
      "subfield": "molecular-biology"
    }
  ],
  "title": "Decoding common and complex genotypes of facioscapulohumeral muscular dystrophy using methylation profiling and long-read sequencing.",
  "title_text": "Decoding common and complex genotypes of facioscapulohumeral muscular dystrophy using methylation profiling and long-read sequencing.",
  "type": "peer-reviewed",
  "updated_at": "2026-10-30T12:20:10Z",
  "url": "https://research.draconitic.com/paper/dk1sav6rkah04",
  "venue": "Brain communications",
  "versions": []
}