title: Myogenic enhancers regulate expression of the facioscapulohumeral muscular dystrophy-associated DUX4 gene.
abstract: Facioscapulohumeral muscular dystrophy (FSHD) is linked to epigenetic dysregulation of the chromosome 4q35 D4Z4 macrosatellite. However, this does not account for the tissue specificity of FSHD pathology, which requires stable expression of an alternative full-length mRNA splice form of DUX4 (DUX4-fl) from the D4Z4 array in skeletal muscle. Here, we describe the identification of two enhancers, DUX4 myogenic enhancer 1 (DME1) and DME2 which activate DUX4-fl expression in skeletal myocytes but not fibroblasts. Analysis of the chromatin revealed histone modifications and RNA polymerase II occupancy consistent with DME1 and DME2 being functional enhancers. Chromosome conformation capture analysis confirmed association of DME1 and DME2 with the DUX4 promoter in vivo. The strong interaction between DME2 and the DUX4 promoter in both FSHD and unaffected primary myocytes was greatly reduced in fibroblasts, suggesting a muscle-specific interaction. Nucleosome occupancy and methylome sequencing analysis indicated that in most FSHD myocytes, both enhancers are associated with nucleosomes but have hypomethylated DNA, consistent with a permissive transcriptional state, sporadic occupancy, and the observed DUX4 expression in rare myonuclei. Our data support a model in which these myogenic enhancers associate with the DUX4 promoter in skeletal myocytes and activate transcription when epigenetically derepressed in FSHD, resulting in the pathological misexpression of DUX4-fl.
authors.affiliation: Wellstone Program, Departments of Cell and Developmental Biology and Neurology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Charis L
authors.initials: CL
authors.lastname: Himeda
authors.forename: Céline
authors.initials: C
authors.lastname: Debarnot
authors.forename: Sachiko
authors.initials: S
authors.lastname: Homma
authors.forename: Mary Lou
authors.initials: ML
authors.lastname: Beermann
authors.forename: Jeffrey B
authors.initials: JB
authors.lastname: Miller
authors.forename: Peter L
authors.initials: PL
authors.lastname: Jones
authors.forename: Takako I
authors.initials: TI
authors.lastname: Jones
mesh_terms: D017398:Alternative Splicing / Q000235:genetics; D002478:Cells, Cultured; D019175:DNA Methylation; D004742:Enhancer Elements, Genetic; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D006657:Histones / Q000235:genetics; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D024510:Muscle Development / Q000235:genetics*; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000378:metabolism; D020391:Muscular Dystrophy, Facioscapulohumeral / Q000235:genetics*; D009707:Nucleosomes / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D012319:RNA Polymerase II / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics
chemical_list: C121257:DUX4L1 protein, human; D006657:Histones; D018398:Homeodomain Proteins; D009707:Nucleosomes; D012333:RNA, Messenger; D012319:RNA Polymerase II