RNA Interference Inhibits DUX4-induced Muscle Toxicity In Vivo: Implications for a Targeted FSHD Therapy

Wallace, Lindsay; Liu, Jian; Domire, Jacqueline S.; Garwick-Coppens, Sara E; Guckes, S.; Mendell, Jerry R.; Flanigan, Kevin M.; Harper, Scott Q.

doi:10.1038/mt.2012.68

Cited by 107 publications

(115 citation statements)

References 32 publications

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“…In healthy individuals with a 'full length' repeat array (410 copies) of D4Z4, the transcription of DUX4 is suppressed by a condensed heterochromatin structure of the surrounding genomic region. [20][21][22] For FSHD1, the second genetic component required is a contraction of D4Z4 repeats to less than 10 units. 10 This is associated with hypomethylation of the D4Z4 repeat array and a relaxation of the chromatin structure allowing for transcription of DUX4.…”

Section: Introductionmentioning

confidence: 99%

Diagnostic approach for FSHD revisited: SMCHD1 mutations cause FSHD2 and act as modifiers of disease severity in FSHD1

et al. 2014

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Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant muscular disorder with a wide clinical variability. Contractions of the D4Z4 macrosatellite repeat on chromosome 4q35 are the molecular basis of the pathophysiology. Recently, in a subset of patients without D4Z4 repeat contractions, variants in the SMCHD1 gene have been identified that lead to hypomethylation of D4Z4 and thus DUX4 transcription, which causes FSHD type 2. In this study, we have screened 55 FSHD1-negative and 40 FSHD1-positive patients from unrelated families for potentially pathogenic variants in SMCHD1 by nextgeneration sequencing (NGS). We identified variants in SMCHD1 in 11 index patients, including missense, splice site and nonsense mutations. We developed a pyrosequencing assay to determine the methylation status of the D4Z4 repeat array and found significantly lower methylation levels for FSHD2 patients than for healthy controls and FSHD1 patients. Two out of eleven SMCHD1 mutation carriers had moderately contracted D4Z4 alleles thus these patients are suffering from FSHD1 and 2. Comparing the phenotype of patients, all FSHD2 patients were relatively mildly affected while patients with FSHD1+2 were much more severely affected than expected from their D4Z4 copy number. Our findings confirm the role of SMCHD1 mutations in FSHD2 and as a modifier of disease severity. With SMCHD1 variants found in 16.4% of phenotypic FSHD patients without D4Z4 repeat contractions, the incidence of FSHD2 is rather high and hence we suggest including sequencing of SMCHD1, haplotyping and methylation analysis in the workflow of molecular FSHD diagnostics.

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Section: Introductionmentioning

confidence: 99%

Diagnostic approach for FSHD revisited: SMCHD1 mutations cause FSHD2 and act as modifiers of disease severity in FSHD1

et al. 2014

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“…Hence, DUX4 are occasionally expressed in skeletal muscle nuclei (14). (32). The study illustrates a 90% reduction in DUX4 protein and 64% reduction in DUX4 mRNA.…”

Section: Epigenetics Of Fshdmentioning

confidence: 69%

“…The basis for anti-inflammatory biologics is to suppress inflammation commonly seen in muscle pathology of FSHD patients in order to slow phenotype progression. All the while, gene therapy has been explored to reduce pathogenic DUX4 protein production in FSHD by controlling D4Z4 methylation, suppressing DUX4 mRNA, and inhibiting DUX4 pathway (5,17,(30)(31)(32). Several inhibitory tools are available for use including small interfering RNA (siRNA), small hairpin RNA (shRNA), microRNA (miRNA) and antisense oligonucleotides (33,34).…”

Section: Therapeutic Approaches To Fshdmentioning

confidence: 99%

Targeting mRNA for the treatment of facioscapulohumeral muscular dystrophy

Bao

Maruyama

Yokota

2016

IRDR

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“…Further, it is still unclear whether there is any sequence polymorphism in the DUX4 gene or flanking regions, as it is difficult to sequence the gene and the DUX4 transcript, which is expressed at the very low levels even in the muscle tissues of patients 14,17 . Since therapeutic approaches including nucleic acid drugs targeting DUX4 mRNA are being studied 18,19 , it may be useful to determine the exact DUX4 sequence of patients for the development of effective therapies as well as an integrative diagnostic method.…”

Section: Discussionmentioning

confidence: 99%

Nanopore-based single molecule sequencing for D4Z4 array responsible for facioscapulohumeral muscular dystrophy

Mitsuhashi

Ueda

et al. 2017

Journal of the Neurological Sciences

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Subtelomeric macrosatellite repeats are difficult to sequence using conventional sequencing methods owing to the high similarity among repeat units and high GC content. Sequencing these repetitive regions is challenging, even with recent improvements in sequencing technologies. Among these repeats, a haplotype carrying a particular sequence and shortening of the D4Z4 array on human chromosome 4q35 causes one of the most prevalent forms of muscular dystrophy with autosomaldominant inheritance, facioscapulohumeral muscular dystrophy (FSHD). Here, we applied a nanoporebased ultra-long read sequencer to sequence a BAC clone containing 13 D4Z4 repeats and flanking regions. We successfully obtained the whole D4Z4 repeat sequence, including the pathogenic gene DUX4 in the last D4Z4 repeat. The estimated sequence accuracy of the total repeat region was 99.8% based on a comparison with the reference sequence. Errors were typically observed between purine or between pyrimidine bases. Further, we analyzed the D4Z4 sequence from publicly available ultra-long whole human genome sequencing data obtained by nanopore sequencing. This technology may be a new tool for studying D4Z4 repeats and pathomechanism of FSHD in the future and has the potential to widen our understanding of subtelomeric regions.Facioscapulohumeral muscular dystrophy (FSHD) is one of the most prevalent adult-onset muscular dystrophies. The genomes of most patients with FSHD have a common feature, i.e., a contracted subtelomeric macrosatellite repeat array called D4Z4 on chromosome 4q35. The D4Z4 array consists of a highly similar 3.3-kb single repeat unit. Normally, the D4Z4 array is highly methylated and forms heterochromatin. Patients with FSHD have less than 11 D4Z4 repeats 1-3 . In Japan, the majority of patients with FSHD have less than 7 repeats 4 . Shortening of the D4Z4 array causes the de-repression of the flanking genes as well as DUX4, located in the last D4Z4 repeat. The ectopic expression of DUX4 is toxic in muscle tissues and is thought to be a causal factor for FSHD [5][6][7][8][9] . In addition to the repeat number, the haplotype of the last D4Z4 repeat is important for the development of FSHD 1,2 . The telomeric flanking region of D4Z4 contains the 3′ UTR of DUX4 and is called the pLAM region. The presence of a polyadenylation signal in this region allows DUX4 expression and disease manifestation 10 . In contrast, individuals without polyadenylation signals do not manifest the disease 2 .

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RNA Interference Inhibits DUX4-induced Muscle Toxicity In Vivo: Implications for a Targeted FSHD Therapy

Cited by 107 publications

References 32 publications

Diagnostic approach for FSHD revisited: SMCHD1 mutations cause FSHD2 and act as modifiers of disease severity in FSHD1

Diagnostic approach for FSHD revisited: SMCHD1 mutations cause FSHD2 and act as modifiers of disease severity in FSHD1

Targeting mRNA for the treatment of facioscapulohumeral muscular dystrophy

Nanopore-based single molecule sequencing for D4Z4 array responsible for facioscapulohumeral muscular dystrophy

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