Facioscapulohumeral muscular dystrophy (FSHD), a common hereditary myopathy, is caused either by the contraction of the D4Z4 macrosatellite repeat at the distal end of chromosome 4q to a size of 1 to ...10 repeat units (FSHD1) or by mutations in D4Z4 chromatin modifiers such as Structural Maintenance of Chromosomes Hinge Domain Containing 1 (FSHD2). These two genotypes share a phenotype characterized by progressive and often asymmetric muscle weakening and atrophy, and common epigenetic alterations of the D4Z4 repeat. All together, these epigenetic changes converge the two genetic forms into one disease and explain the derepression of the DUX4 gene, which is otherwise kept epigenetically silent in skeletal muscle. DUX4 is consistently transcriptionally upregulated in FSHD1 and FSHD2 skeletal muscle cells where it is believed to exercise a toxic effect. Here we provide a review of the recent literature describing the progress in understanding the complex genetic and epigenetic architecture of FSHD, with a focus on one of the consequences that these epigenetic changes inflict, the DUX4‐induced immune deregulation cascade. Moreover, we review the latest therapeutic strategies, with particular attention to the potential of epigenetic correction of the FSHD locus.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Immunoglobulin 4 (IgG4) is one of four human IgG subclasses and has several unique functional characteristics. It exhibits low affinity for complement and for most Fc receptors. It furthermore has ...generally high affinity for its antigen, with binding occurring in a monovalent fashion, as IgG4 can exchange Fab‐arms with other IgG4 molecules. Because of these characteristics, IgG4 is believed to block its targets and prevent inflammation, which, depending on the setting, can have a protective or pathogenic effect. One example of IgG4 pathogenicity is muscle‐specific kinase (MuSK) myasthenia gravis (MG), in which patients develop IgG4 MuSK autoantibodies, resulting in muscle weakness. As a consequence of the distinct IgG4 characteristics, the pathomechanism of MuSK MG is very different from IgG1‐and IgG3‐mediated autoimmune diseases, such as acetylcholine receptor MG. In recent years, new autoantibodies in a spectrum of autoimmune diseases have been discovered. Interestingly, some were found to be predominantly IgG4. These IgG4‐mediated autoimmune diseases share many pathomechanistic aspects with MuSK MG, suggesting that IgG4‐mediated autoimmunity forms a separate niche among the antibody‐mediated disorders. In this review, we summarize the group of IgG4‐mediated autoimmune diseases, discuss the role of IgG4 in MuSK MG, and highlight interesting future research questions for IgG4‐mediated autoimmunity.
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BFBNIB, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK
The genetic diagnostics of inherited neuromuscular diseases (NMDs) is challenging due to their clinical and genetic heterogeneity. We launched an online survey within the EURO-NMD European Reference ...Network (ERN) to collect information about the availability/distribution of genetic testing across 61 ERN health care providers (HCPs). A 17 items questionnaire was designed to address methods used, the number of genetic tests available, the clinical pathway to access genetic testing, the use of next-generation sequencing (NGS) and participation to quality assessment schemes (QAs). A remarkable number of HCPs (49%) offers greater than or equal to 500 genetic tests per year, 43,6% offers 100-500 genetic tests per year, and 7,2% less than or equal to 100 per year. NGS is used by 94% of centres, Sanger sequencing by 84%, MLPA by 66% and Southern blotting by 36%. The majority of centres (60%) offer NGS for all patients that fulfil criteria for NMD of genetic origin. Pipelines for NGS vary amongst centres, even within the same national system. Referral of patients to genetic laboratories by specialists was frequently reported (58%), and 65% of centres participates in genetic testing QAs. We specifically evaluated how many centres cover SMA, DMD, Pompe, LGMDs, and TTR genes/diseases genetic diagnosis, since these rare diseases benefit from personalised therapies. We used the Orphanet EUGT numbers, provided by 82% of HCPs. SMA, DMD, LGMD, TTR and GAA genes are covered by EUGTs although with different numbers and modalities. The number of genetic tests for NMDs offered across HCPs National Health systems is quite high, including routine techniques and NGS. The number and type of tests offered and the clinical practices differ among centres. We provided evidence that survey tools might be useful to learn about the state-of-the-art of ERN health-related activities and to foster harmonisation and standardisation of the complex care for the rare disease patients in the EU.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Each unit of the D4Z4 macrosatellite repeat contains a retrotransposed gene encoding the DUX4 double-homeobox transcription factor. Facioscapulohumeral dystrophy (FSHD) is caused by deletion of a ...subset of the D4Z4 units in the subtelomeric region of chromosome 4. Although it has been reported that the deletion of D4Z4 units induces the pathological expression of DUX4 mRNA, the association of DUX4 mRNA expression with FSHD has not been rigorously investigated, nor has any human tissue been identified that normally expresses DUX4 mRNA or protein. We show that FSHD muscle expresses a different splice form of DUX4 mRNA compared to control muscle. Control muscle produces low amounts of a splice form of DUX4 encoding only the amino-terminal portion of DUX4. FSHD muscle produces low amounts of a DUX4 mRNA that encodes the full-length DUX4 protein. The low abundance of full-length DUX4 mRNA in FSHD muscle cells represents a small subset of nuclei producing a relatively high abundance of DUX4 mRNA and protein. In contrast to control skeletal muscle and most other somatic tissues, full-length DUX4 transcript and protein is expressed at relatively abundant levels in human testis, most likely in the germ-line cells. Induced pluripotent (iPS) cells also express full-length DUX4 and differentiation of control iPS cells to embryoid bodies suppresses expression of full-length DUX4, whereas expression of full-length DUX4 persists in differentiated FSHD iPS cells. Together, these findings indicate that full-length DUX4 is normally expressed at specific developmental stages and is suppressed in most somatic tissues. The contraction of the D4Z4 repeat in FSHD results in a less efficient suppression of the full-length DUX4 mRNA in skeletal muscle cells. Therefore, FSHD represents the first human disease to be associated with the incomplete developmental silencing of a retrogene array normally expressed early in development.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
In the majority of facioscapulohumeral muscular dystrophy (FSHD) cases, the molecular basis of the disease is due to loss of subtelomeric D4Z4 repeat units at 4q35. Occasionally, an apparent absence ...of the contracted D4Z4 repeat is associated with FSHD. One explanation for this finding is a deletion in the region proximal to the D4Z4 repeat array that encompasses the p13E-11 (D4F104S1) probe-binding site used in the DNA diagnosis. The frequency of such proximally extended deletions is unknown, and to date, few patients have been described due to the difficulties in the molecular identification of such cases.
We describe a family (DUK 2531) in which a contracted D4Z4 allele and a large proximal deletion of approximately 75 kb are segregating to 11 individuals. This is the largest deletion identified to date. Family DUK 2531 was initially thought to have normal D4Z4 fragment size and therefore unlinked to the 4q35 region (FSHD1B).
Further molecular analysis of DUK 2531 reveals the presence of 10 repeat units (33 kb). The extended deletion includes the probe p13E-11 and B31 binding sites, the inverted repeat D4S2463, and genes FRG2 and TUBB4Q.
Despite the length of the proximal deletion in this family, the range and severity of the clinical manifestations are typical for the disorder. Because such deletions can lead to misinterpretation in the diagnostic setting, this suggests the need for additional diagnostic tests in facioscapulohumeral muscular dystrophy.
Facioscapulohumeral muscular dystrophy type 1 (FSHD1) is caused by contraction of the D4Z4 repeat array on chromosome 4 to a size of 1–10 units. The residual number of D4Z4 units inversely correlates ...with clinical severity, but significant clinical variability exists. Each unit contains a copy of the DUX4 retrogene. Repeat contractions are associated with changes in D4Z4 chromatin structure that increase the likelihood of DUX4 expression in skeletal muscle, but only when the repeat resides in a genetic background that contains a DUX4 polyadenylation signal. Mutations in the structural maintenance of chromosomes flexible hinge domain containing 1 (SMCHD1) gene, encoding a chromatin modifier of D4Z4, also result in the increased likelihood of DUX4 expression in individuals with a rare form of FSHD (FSHD2). Because SMCHD1 directly binds to D4Z4 and suppresses somatic expression of DUX4, we hypothesized that SMCHD1 may act as a genetic modifier in FSHD1. We describe three unrelated individuals with FSHD1 presenting an unusual high clinical severity based on their upper-sized FSHD1 repeat array of nine units. Each of these individuals also carries a mutation in the SMCHD1 gene. Familial carriers of the FSHD1 allele without the SMCHD1 mutation were only mildly affected, suggesting a modifier effect of the SMCHD1 mutation. Knocking down SMCHD1 in FSHD1 myotubes increased DUX4 expression, lending molecular support to a modifier role for SMCHD1 in FSHD1. We conclude that FSHD1 and FSHD2 share a common pathophysiological pathway in which the FSHD2 gene can act as modifier for disease severity in families affected by FSHD1.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Myasthenia gravis (MG) is a severely debilitating autoimmune disease that is due to a decrease in the efficiency of synaptic transmission at neuromuscular synapses. MG is caused by antibodies against ...postsynaptic proteins, including (i) acetylcholine receptors, the neurotransmitter receptor, (ii) muscle-specific kinase (MuSK), a receptor tyrosine kinase essential for the formation and maintenance of neuromuscular synapses, and (iii) low-density lipoprotein receptor-related protein 4 (Lrp4), which responds to neural Agrin by binding and stimulating MuSK. Passive transfer studies in mice have shown that IgG4 antibodies from MuSK MG patients cause disease without requiring complement or other immune components, suggesting that these MuSK antibodies cause disease by directly interfering with MuSK function. Here we show that pathogenic IgG4 antibodies to MuSK bind to a structural epitope in the first Ig-like domain of MuSK, prevent binding between MuSK and Lrp4, and inhibit Agrin-stimulated MuSK phosphorylation. In contrast, these IgG4 antibodies have no direct effect on MuSK dimerization or MuSK internalization. These results provide insight into the unique pathogenesis of MuSK MG and provide clues toward development of specific treatment options.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
The human double-homeodomain retrogene DUX4 is expressed in the testis and epigenetically repressed in somatic tissues. Facioscapulohumeral muscular dystrophy (FSHD) is caused by mutations that ...decrease the epigenetic repression of DUX4 in somatic tissues and result in mis-expression of this transcription factor in skeletal muscle. DUX4 binds sites in the human genome that contain a double-homeobox sequence motif, including sites in unique regions of the genome as well as many sites in repetitive elements. Using ChIP-seq and RNA-seq on myoblasts transduced with DUX4 we show that DUX4 binds and activates transcription of mammalian apparent LTR-retrotransposons (MaLRs), endogenous retrovirus (ERVL and ERVK) elements, and pericentromeric satellite HSATII sequences. Some DUX4-activated MaLR and ERV elements create novel promoters for genes, long non-coding RNAs, and antisense transcripts. Many of these novel transcripts are expressed in FSHD muscle cells but not control cells, and thus might contribute to FSHD pathology. For example, HEY1, a repressor of myogenesis, is activated by DUX4 through a MaLR promoter. DUX4-bound motifs, including those in repetitive elements, show evolutionary conservation and some repeat-initiated transcripts are expressed in healthy testis, the normal expression site of DUX4, but more rarely in other somatic tissues. Testis expression patterns are known to have evolved rapidly in mammals, but the mechanisms behind this rapid change have not yet been identified: our results suggest that mobilization of MaLR and ERV elements during mammalian evolution altered germline gene expression patterns through transcriptional activation by DUX4. Our findings demonstrate a role for DUX4 and repetitive elements in mammalian germline evolution and in FSHD muscular dystrophy.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The neuromuscular disorder facioscapulohumeral muscular dystrophy (FSHD) results from integral deletions of the subtelomeric repeat D4Z4 on chromosome 4q. A disruption of chromatin structure ...affecting gene expression is thought to underlie the pathophysiology. The global gene expression profiling of mature muscle tissue presented here provides the first insight into an FSHD-specific defect in myogenic differentiation. FSHD expression profiles generated by oligonucleotide microarrays were compared with those from normal muscle as well as other types of muscular dystrophies (DMD, aSGD) in order to determine FSHD-specific changes. In addition, matched biopsies (affected and unaffected muscle) from individuals with FSHD served to monitor expression changes during the progression of the disease as well as to diminish non-specific changes resulting from individual variability. Among genes altered in an FSHD-specific and highly significant manner, many are involved in myogenic differentiation and suggest a partial block in the normal differentiation program. Indeed, many of the transcripts affected in FSHD represent direct targets of the transcription factor MyoD. Additional mis-expressed genes confirm a diminished capacity to buffer oxidative stress, as demonstrated in FSHD myoblasts. This enhanced vulnerability of proliferative stage myoblasts to reactive oxygen species is also disease-specific, further implicating a defect in FSHD muscle satellite cells. Importantly, none of the genes localizing to the FSHD region at 4q35 were found to exhibit a significantly altered pattern of expression in FSHD muscle. This finding was corroborated by expression analysis of FSHD muscle using a custom cDNA microarray containing 51 genes and ESTs from the 4q35 region. Disruptions in FSHD myogenesis and oxidative capacity may therefore not arise from a position effect mechanism as has been previously suggested, but rather from a global effect on gene regulation. Improper nuclear localization of 4qter is discussed as an alternative model for FSHD gene regulation and pathogenesis.
In mammalian embryos, proper zygotic genome activation (ZGA) underlies totipotent development. Double homeobox (DUX)-family factors participate in ZGA, and mouse Dux is required for forming cultured ...two-cell (2C)-like cells. Remarkably, in mouse embryonic stem cells, Dux is activated by the tumor suppressor p53, and Dux expression promotes differentiation into expanded-fate cell types. Long-read sequencing and assembly of the mouse Dux locus reveals its complex chromatin regulation including putative positive and negative feedback loops. We show that the p53-DUX/DUX4 regulatory axis is conserved in humans. Furthermore, we demonstrate that cells derived from patients with facioscapulohumeral muscular dystrophy (FSHD) activate human DUX4 during p53 signaling via a p53-binding site in a primate-specific subtelomeric long terminal repeat (LTR)10C element. In summary, our work shows that p53 activation convergently evolved to couple p53 to Dux/DUX4 activation in embryonic stem cells, embryos and cells from patients with FSHD, potentially uniting the developmental and disease regulation of DUX-family factors and identifying evidence-based therapeutic opportunities for FSHD.
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GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ
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