**8. Conclusions**

40 Neuromuscular Disorders

complex in vitro and in vivo comprising of YY1, HMGB2 and nucleolin, termed D4Z4 Recognition Complex (DRC) (Gabellini et al., 2002). The ubiquitous transcription factor Yin Yang 1 (YY1) is a recruiter of polycomb group proteins (PcG), which are responsible for chromatin remodelling and epigenetic silencing in many fundamental biological processes. YY1, exerts its effects on genes involved in normal biologic processes such as embryogenesis, differentiation, replication, and cellular proliferation. Its ability to initiate, activate, or repress transcription depends upon context (Gordon et al., 2006). Furthermore, the activity of YY1 is modulated by histone deacetylases and histone acetyltransferases (Yao et al., 2001). HMGB2 is a member of one of the three families of high mobility group (HMG) proteins (Bustin, 1999; Bianchi and Beltrame, 2000;Agresti and Bianchi, 2003). It has been proposed that HMGB2 might be involved in the organization and/or maintenance of heterochromatic regions through the SP100-mediated interaction with HP1 (Lehming et al., 1998). The third component of the DRC, nucleolin, is an abundant nucleolar protein, which has been implicated in chromatin structure, ribosomal RNA (rRNA) transcription, rRNA maturation, ribosome assembly and nucleo-cytoplasmic transport. To address whether the level of the DRC components influenced transcription of 4q35 genes, antisense experiments to decrease intracellular levels of DRC components were performed. These experiments showed that depletion of YY1, HMGB2 or nucleolin results in overexpression of the 4q35 gene *FRG2*, which is silent in normal cells and tissues (Gabellini et al., 2002). Accumulating evidences indicate that gene regulation can be affected by physical interaction between two distant chromosomal regions in *cis* and in *trans* in mammalian cells (Tolhuis et al., 2002; Horike et al., 2005; Spilianakis et al., 2005; Lomvardas et al., 2006). Thus the DRC might exerts is inhibitory activity either modifying the chromatin structure or acting directly on 4q35 genes promoters through a physical interaction mediated by the formation of a cromatin loop (Gabellini et al., 2002; Pirozhkova et al., 2008). The physical interaction between D4Z4 and FRG1 has been demonstrated (Pirozhkova et al., 2008; Bodega et al., 2009) in normal myoblast by Chromosome conformation capture (3C), which is a technique that identifies long distance intra- and inter-chromosomal interactions (Dekker et al., 2002). Interestingly chromatin seems to undergo remodeling during myogenic differentiation. It has been shown that in normal myoblasts, the *FRG1* gene is repressed and its promoter physically interacts with the D4Z4 array; upon differentiation, *FRG1* gene is expressed and the chromatin loop between FRG1 promoter and D4Z4 is relaxed (Bodega et al.2009). Consistent with the observed mis-regulation of FRG1, a small reduction in the D4Z4–FRG1 promoter interaction was observed in FSHD myoblasts compared with controls (Bodega et al., 2009). Different findings obtained with 3C analysis described the formation of loops between other elements in the FSHD locus (DUX4c and the 4qA/B marker) and the FRG1 promoter (Pirozhkova et al., 2008). These data indicate that the tridimensional structure of the FSHD locus is complex and composite, probably more than one sequence elements (for example, D4Z4, DUX4c,4qA/B) or more than one chromatin modification factor might be required to obtain a fine regulation of *FRG1* gene expression during muscle differentiation

The nucleoplasm is a high defined and structured compartment and chromosomes occupy specific and distinct territories. These chromosome territories are related to gene density,

(Petrov et al., 2006; Pirozhkova et al., 2008).

**7.4 Subnuclear localization of 4q35** 

D4Z4 repeat contraction in patients with FSHD was discovered almost 20 years ago, nevertheless the exact molecular mechanism causing the FSHD phenotype has still not been elucidated and the search for a unifying model that can explain all the clinical features that have been observed in time has been frustrated. No histological or biochemical markers are available to independently confirm a specific FSHD diagnosis that remains mainly clinical. The molecular test primarily used for FSHD diagnosis was based on the initial observation that 95% of FSHD patients carry a reduction of integral numbers of D4Z4 repeats at 4q35 with full penetrance (Van Deutekom et al., 2003). However the wide use of this test revealed several exceptions to the original assumption. Through the years the threshold size of D4Z4 alleles has been increased from the original 28 kb (6 D4Z4 repeats) (Wimenga et al., 1999b) to 35 kb (8 D4Z4 repeats) (Van Deutekom et al., 2003), with FSHD cases carrying D4Z4 alleles of 38-41 kb (9-11 D4Z4 repeats) considered borderline alleles (Butz et al., 2003; Vitelli et al., 1999). A further analysis of genotype-phenotype correlation led in time to the identification of subjects carrying D4Z4 reduced alleles with no sign of muscle weakness in FSHD families (Ricci et al., 1999; Tonini et al., 2004) as well as in normal controls (Van Overveld et al., 2000; Weiffenbach et al., 1992). The genotype-phenotype correlation conducted more recently on a large scale using a standardized method of evaluation allowed to estimate that 1) 20% of FSHD patients carry full-length D4Z4 alleles, 2) over 25% of relatives carrying D4Z4 reduced alleles do not have FSHD, 3) 3% of healthy subjects from the general population carry D4Z4 reduced alleles 4) no specific 4q haplotype is uniquely associated with FSHD. Remarkably, these studies established as a general rule rather than an exception that detection of a D4Z4 reduced allele is not sufficient to predict FSHD (Scionti et al., 2012a; Scionti et al., 2012b). Over the years, the molecular etiology of FSHD

Facioscapulohumeral Muscular Dystrophy: From Clinical Data to Molecular Genetics and Return 43

Bannister, A.J., Zegerman, P., Partridge, J.F., Miska, E.A., Thomas, J.O., Allshire, R.C., &

Bauer, M.K., Schubert, A., Rocks, O., & Grimm, S. (1999). Adenine nucleotide translocase-1,

Bernard, P., Maure, J.F., Partridge, J.F., Genier, S., Javerzat, J.P., & Allshire, R.C. (2001).

Bernstein, B.E., Meissner, A., & Lander, E.S. (2007). The mammalian epigenome. *Cell* 128,

Bessonov, S., Anokhina, M., Krasauskas, A., Golas, M.M., Sander, B., Will, C.L., Urlaub, H.,

Bianchi M. E. & Beltrame M. (2000) Upwardly mobile proteins. Workshop: the role of HMG

Bodega, B., Ramirez, G.D., Grasser, F., Cheli, S., Brunelli, S., Mora, M., Meneveri, R.,

Bortolanza, S., Nonis, A., Sanvito, F., Maciotta S., Sitia, G., Wei, J., Torrente, Y., Di Serio, C.,

Bosnakovski, D., Xu, Z., Gang, E.J., Galindo, C.L., Liu, M., Simsek, T., Garner, H.R., Agha-

Brouwer, O.F., Padberg, G.W., van der Ploeg, R.J., Ruys, C.J., & Brand, R. (1992). The

Celegato, B., Capitanio, D., Pescatori, M., Romualdi, C., Pacchioni, B., Cagnin, S., Vigano, A.,

Cheli, S., Francois, S., Bodega, B., Ferrari, F., Tenedini, E., Roncaglia, E., Ferrari, S., Ginelli,

of MyoD-dependent genes. *Proteomics* 6, 5303-5321.

facioscapulohumeral muscular dystrophy. *Brain* 115 ( Pt 5), 1587-1598. Bustin M. (1999) Regulation of DNA-dependent activities by the functional motifs of the High-Mobility-Group chromosomal proteins. *Mol. Cell. Biol*. 19: 5237–5246 Butz, M., Koch, M.C., Muller-Felber, W., Lemmers, R.J., van der Maarel, S.M., & Schreiber,

spliceosome activation and first step catalysis. *RNA* 16, 2384-2403.

the HP1 chromo domain. *Nature* 410, 120-124.

*J Cell Biol 147*, 1493-1502.

differentiation. *BMC Biol* 7, 41.

pathologies. *EMBO J*. *27*, 2766-2779.

*Mol Ther.* 11:2055-64

2539-2542.

669-681.

109–114

Kouzarides, T. (2001). Selective recognition of methylated lysine 9 on histone H3 by

a component of the permeability transition pore, can dominantly induce apoptosis.

Requirement of heterochromatin for cohesion at centromeres. *Science* 294,

Stark, H., & Luhrmann, R. (2010). Characterization of purified human Bact spliceosomal complexes reveals compositional and morphological changes during

proteins in chromatin structure,gene expression and neoplasia. *EMBO Rep*. 1:

Marozzi, A., Mueller, S., Battaglioli, E., *et al.* (2009). Remodeling of the chromatin structure of the facioscapulohumeral muscular dystrophy (FSHD) locus and upregulation of FSHD-related gene 1 (FRG1) expression during human myogenic

Chamberlain, J.R. & Gabellini D. (2011). AAV6-mediated systemic shRNA delivery reverses disease in a mouse model of facioscapulohumeral muscular dystrophy.

Mohammadi, S., Tassin, A., Coppee, F.*, et al.* (2008). An isogenetic myoblast expression screen identifies DUX4-mediated FSHD-associated molecular

influence of handedness on the distribution of muscular weakness of the arm in

H. (2003). Facioscapulohumeral muscular dystrophy. Phenotype-genotype correlation in patients with borderline D4Z4 repeat numbers. *J Neurol* 250, 932-937.

Colantoni, L., Begum, S., Ricci, E., *et al.* (2006). Parallel protein and transcript profiles of FSHD patient muscles correlate to the D4Z4 arrangement and reveal a common impairment of slow to fast fibre differentiation and a general deregulation

E., & Meneveri, R. (2011). Expression Profiling of FSHD-1 and FSHD-2 Cells during

has remained enigmatic, and the literature is filled with claims of causes that fail to be confirmed by other groups. Indeed, this might be expected if the clinical manifestation of FSHD symptoms is not only dependent on the structure/haplotype of D4Z4 contractions. This does not exclude an important pathogenic role for DUX4 or other candidate factors, but do establish a complex mechanism beyond current understanding indicating that a profound re-thinking of the genetic disease mechanism and modes of inheritance of FSHD is required.

In-depth examination of disease points to a more complex genetic etiology in which D4Z4 reduction might play a significant role only in association with other determinants, including genetic, epigenetic and environmental factors. Indeed, it is possible that in the heterozygous state a D4Z4 reduction might produce a predisposing condition that requires other epigenetic mechanisms or mutations in additional genes, both in *cis* and in *trans,* to cause overt myopathy. Finally it is also plausible that drugs or toxic agents might contribute to the disease onset and clinical variability. It is likely that, all mechanisms described above may contribute to the diverse phenotypic expression observed in carrier of D4Z4 reduced alleles. One of the major challenges for clinicians and researchers involved in FSHD studies will be to establish the weight that each single factor has in FSHD development. Particular attention should be paid to the relevance of epigenetics in the pathogenesis of FSHD. At the 4q subtelomere chromatin is normally tightly packed, probably as facultative heterochromatin. However this region can be highly dynamic as demonstrated by the fact that in patients with FSHD, this chromatin structure becomes more open. As a consequence, regulation of candidate genes can be influenced by proteins that may bind to or be released from D4Z4. One of the major goals for future FSHD research will be to integrate these disease mechanisms into a single model that can be used to explain the clinical data and to improve the molecular diagnosis; both steps are essential to develop effective therapeutic strategies.

#### **9. References**


has remained enigmatic, and the literature is filled with claims of causes that fail to be confirmed by other groups. Indeed, this might be expected if the clinical manifestation of FSHD symptoms is not only dependent on the structure/haplotype of D4Z4 contractions. This does not exclude an important pathogenic role for DUX4 or other candidate factors, but do establish a complex mechanism beyond current understanding indicating that a profound re-thinking of the genetic disease mechanism and modes of inheritance of FSHD is

In-depth examination of disease points to a more complex genetic etiology in which D4Z4 reduction might play a significant role only in association with other determinants, including genetic, epigenetic and environmental factors. Indeed, it is possible that in the heterozygous state a D4Z4 reduction might produce a predisposing condition that requires other epigenetic mechanisms or mutations in additional genes, both in *cis* and in *trans,* to cause overt myopathy. Finally it is also plausible that drugs or toxic agents might contribute to the disease onset and clinical variability. It is likely that, all mechanisms described above may contribute to the diverse phenotypic expression observed in carrier of D4Z4 reduced alleles. One of the major challenges for clinicians and researchers involved in FSHD studies will be to establish the weight that each single factor has in FSHD development. Particular attention should be paid to the relevance of epigenetics in the pathogenesis of FSHD. At the 4q subtelomere chromatin is normally tightly packed, probably as facultative heterochromatin. However this region can be highly dynamic as demonstrated by the fact that in patients with FSHD, this chromatin structure becomes more open. As a consequence, regulation of candidate genes can be influenced by proteins that may bind to or be released from D4Z4. One of the major goals for future FSHD research will be to integrate these disease mechanisms into a single model that can be used to explain the clinical data and to improve the molecular diagnosis; both steps are essential to develop effective therapeutic

Agresti A. & Bianchi M. E. (2003) HMGB proteins and gene expression. *Curr. Opin. Genet.* 

Amrichova, J., Lukasova, E., Kozubek, S., & Kozubek, M. (2003). Nuclear and territorial

Arashiro, P., Eisenberg, I., Kho, A.T., Cerqueira, A.M., Canovas, M., Silva, H.C., Pavanello,

Bakker, E., Van der Wielen, M.J., Voorhoeve, E., Ippel, P.F., Padberg, G.W., Frants, R.R., &

Bakker, E., Wijmenga, C., Vossen, R.H., Padberg, G.W., Hewitt, J., van der Wielen, M.,

on 4q35 has a homologue on 10qter. *Muscle Nerve* 2, S39-44.

topography of chromosome telomeres in human lymphocytes. *Exp Cell Res 289*, 11-26.

R.C., Verjovski-Almeida, S., Kunkel, L.M., & Zatz, M. (2009). Transcriptional regulation differs in affected facioscapulohumeral muscular dystrophy patients compared to asymptomatic related carriers. *Proc Natl Acad Sci U S A* 106, 6220-6225.

Wijmenga, C. (1996). Diagnostic, predictive, and prenatal testing for facioscapulohumeral muscular dystrophy: diagnostic approach for sporadic and

Rasmussen, K., & Frants, R.R. (1995). The FSHD-linked locus D4F104S1 (p13E-11)

required.

strategies.

**9. References** 

*Dev*. 13: 170–178

familial cases. *J Med Genet* 33, 29-35.


Facioscapulohumeral Muscular Dystrophy: From Clinical Data to Molecular Genetics and Return 45

Fitzsimons, R.B., Gurwin, E.B., & Bird, A.C. (1987). Retinal vascular abnormalities in

Flanigan, K. M. in *Myology* (eds Engel, A. & Franzini-Armstrong, C.) 1123–1133 (McGraw

Frenster, J.H., Allfrey, V.G., & Mirsky, A.E. (1963). Repressed and Active Chromatin Isolated from Interphase Lymphocytes. *Proc Natl Acad Sci U S A* 50, 1026-1032. Gabellini, D., D'Antona, G., Moggio, M., Prelle, A., Zecca, C., Adami, R., Angeletti, B.,

muscular dystrophy in mice overexpressing FRG1. *Nature* 439, 973-977. Gabellini, D., Green, M.R., & Tupler, R. (2002). Inappropriate gene activation in FSHD: a

Gabriels, J., Beckers, M.C., Ding, H., De Vriese, A., Plaisance, S., van der Maarel, S.M.,

Gerace, L., & Burke, B. (1988). Functional organization of the nuclear envelope. *Annu Rev* 

Gordon, S., Akopyan, G., Garban, H., & Bonavida, B. (2006). Transcription factor YY1:

Griggs, R.C., Tawil, R., Storvick, D., Mendell, J.R., & Altherr, M.R. (1993). Genetics of

Hanel, M.L., Sun, C.Y., Jones, T.I., Long, S.W., Zanotti, S., Milner, D., & Jones, P.L. (2011).

Hanel, M.L., Wuebbles, R.D., & Jones, P.L. (2009). Muscular dystrophy candidate gene FRG1

Hark, A.T., Schoenherr, C.J., Katz, D.J., Ingram, R.S., Levorse, J.M., & Tilghman, S.M. (2000).

Hewitt, J.E., Lyle, R., Clark, L.N., Valleley, E.M., Wright, T.J., Wijmenga, C., van Deutekom,

Horike, S., Cai, S., Miyano, M., Cheng, J.F., & Kohwi-Shigematsu, T. (2005). Loss of silent-

Jiang, G., Yang, F., van Overveld, P.G., Vedanarayanan, V., van der Maarel, S., & Ehrlich, M.

dynamic nuclear and sarcomeric protein. *Differentiation* 81, 107-118.

is critical for muscle development. *Dev Dyn* 238, 1502-1512.

subtelomeric 4q. *Hum Mol Genet* 12, 2909-2921.

putative gene within each 3.3 kb element. *Gene* 236, 25-32.

therapeutic implications. *Brain* 110 ( Pt 3), 631-648.

Hill Professional, New York, 2004)

110, 339-348.

*Cell Biol 4*, 335-374.

*Neurology* 43, 2369-2372.

locus. *Nature* 405, 486-489.

*Genet* 3, 1287-1295.

37, 31-40.

1125-1142.

facioscapulohumeral muscular dystrophy. A general association with genetic and

Ciscato, P., Pellegrino, M.A., Bottinelli, R., *et al.* (2006). Facioscapulohumeral

repressor complex binds a chromosomal repeat deleted in dystrophic muscle. *Cell*

Padberg, G.W., Frants, R.R., Hewitt, J.E., Collen, D., *et al.* (1999). Nucleotide sequence of the partially deleted D4Z4 locus in a patient with FSHD identifies a

structure, function, and therapeutic implications in cancer biology. *Oncogene* 25,

facioscapulohumeral muscular dystrophy: new mutations in sporadic cases.

Facioscapulohumeral muscular dystrophy (FSHD) region gene 1 (FRG1) is a

CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2

J.C., Francis, F., Sharpe, P.T., Hofker, M., *et al.* (1994). Analysis of the tandem repeat locus D4Z4 associated with facioscapulohumeral muscular dystrophy. *Hum Mol* 

chromatin looping and impaired imprinting of DLX5 in Rett syndrome. *Nat Genet*

(2003). Testing the position-effect variegation hypothesis for facioscapulohumeral muscular dystrophy by analysis of histone modification and gene expression in

Myogenic Differentiation Evidences Common & Distinctive Gene Dysregulation Patterns. *PLoS One* 6, e20966.


Cheung, P., Allis, C.D., & Sassone-Corsi, P. (2000). Signaling to chromatin through histone

Chuenkongkaew, W.L., Lertrit, P., Limwongse, C., Nilanont, Y., Boonyapisit, K., Sangruchi,

de Greef, J.C., Frants, R.R., & van der Maarel, S.M. (2008). Epigenetic mechanisms of

de Greef, J.C., Lemmers, R.J., van Engelen, B.G., Sacconi, S., Venance, S.L., Frants, R.R.,

Deidda, G., Cacurri, S., Grisanti, P., Vigneti, E., Piazzo, N., & Felicetti, L. (1995). Physical

Deidda, G., Cacurri, S., Piazzo, N., & Felicetti, L. (1996). Direct detection of 4q35

Dekker, J., Rippe, K., Dekker, M., & Kleckner, N. (2002). Capturing chromosome

Dixit, M., Ansseau, E., Tassin, A., Winokur, S., Shi, R., Qian, H., Sauvage, S., Matteotti, C.,

Felice, K.J., & Moore, S.A. (2001). Unusual clinical presentations in patients harboring the facioscapulohumeral dystrophy 4q35 deletion. *Muscle Nerve* 24, 352-356. Felice, K.J., & Whitaker, C.H. (2005). The Clinical Features of Facioscapulohumeral Muscular

Felice, K.J., North, W.A., Moore, S.A., & Mathews, K.D. (2000). FSH dystrophy 4q35 deletion

Figueroa, J.J., & Chapin, J.E. (2010). Isolated facial diplegia and very late-onset myopathy in

Filosto, M., Tonin, P., Scarpelli, M., Savio, C., Greco, F., Mancuso, M., Vattemi, G., Govoni,

Finch, J.T., Lutter, L.C., Rhodes, D., Brown, R.S., Rushton, B., Levitt, M. & Klug, A. (1977) Structure of nucleosome core particle of chromatin. *Nature* 269: 29-36

facioscapulohumeral muscular dystrophy. Mutat Res 647, 94-102.

chromosome 4qter. *Eur J Hum Genet* 3, 155-167.

PITX1. *Proc Natl Acad Sci U S A 104*, 18157-18162.

phenotype. *Neuromuscul Disord* 18, 204-209.

conformation. *Science* 295, 1306-1311.

Patterns. *PLoS One* 6, e20966.

*Neurol* 12, 388-391.

*Med Genet* 33, 361-365.

*Neuromuscul Dis* 6, 119-126.

1927-1931.

444-446.

1449-1459.

modifications. *Cell* 103, 263-271.

Myogenic Differentiation Evidences Common & Distinctive Gene Dysregulation

T., Chirapapaisan, N., & Suphavilai, R. (2005). An unusual family with Leber's hereditary optic neuropathy and facioscapulohumeral muscular dystrophy. *Eur J* 

Tawil, R., & van der Maarel, S.M. (2009). Common epigenetic changes of D4Z4 in contraction-dependent and contraction-independent FSHD. *Hum Mutat* 30,

mapping evidence for a duplicated region on chromosome 10qter showing high homology with the facioscapulohumeral muscular dystrophy locus on

rearrangements implicated in facioscapulohumeral muscular dystrophy (FSHD). *J* 

van Acker, A.M., Leo, O.*, et al.* (2007). DUX4, a candidate gene of facioscapulohumeral muscular dystrophy, encodes a transcriptional activator of

Dystrophy Associated With Borderline (>/=35 kb) 4q35 EcoRI Fragments. *J Clin* 

in patients presenting with facial-sparing scapular myopathy. *Neurology* 54,

two siblings: atypical presentations of facioscapulohumeral dystrophy. *J Neurol* 257,

V., Rizzuto, N., Tupler, R., *et al.* (2008). Novel mitochondrial tRNA Leu(CUN) transition and D4Z4 partial deletion in a patient with a facioscapulohumeral


Facioscapulohumeral Muscular Dystrophy: From Clinical Data to Molecular Genetics and Return 47

Lemmers, R.J., van der Maarel, S.M., van Deutekom, J.C., van der Wielen, M.J., Deidda, G.,

Lemmers, R.J.L., de Kievit, P., van Geel, M., van der Wielen, M.J., Bakker, E., Padberg, G.W.,

Lemmers, R.J., van der Wielen, M.J., Bakker, E., Padberg, G.W., Frants, R.R., & van der

Lemmers, R.J., Wohlgemuth, M., van der Gaag, K.J., van der Vliet, P.J., van Teijlingen, C.M.,

Lemmers, R.J., van der Vliet, P.J., van der Gaag, K.J., Zuniga, S., Frants, R.R., de Knijff, P., &

Levy, S.E., Chen, Y.S., Graham, B.H., & Wallace, D.C. (2000). Expression and sequence

Liu, Q., Jones, T.I., Tang, V.W., Brieher, W.M., & Jones, P.L. (2010). Facioscapulohumeral

Lomvardas, S., Barnea, G., Pisapia, D.J., Mendelsohn, M., Kirkland, J., & Axel, R. (2006). Interchromosomal interactions and olfactory receptor choice. *Cell* 126, 403-413. Luderus, M.E., van Steensel, B., Chong, L., Sibon, O.C., Cremers, F.F., & de Lange, T. (1996).

Lunt, P.W., Jardine, P.E., Koch, M.C., Maynard, J., Osborn, M., Williams, M., Harper, P.S., &

Lunt, P.W. (1998). 44th ENMC International Workshop: Facioscapulohumeral Muscular

Lyle, R., Wright, T.J., Clark, L.N., & Hewitt, J.E. (1995). The FSHD-associated repeat, D4Z4,

human evolution. Am J Hum Genet *86*, 364-377.

with muscle-attachment sites. *J Cell Sci 123*, 1116-1123.

mammalian telomeric complex. *J Cell Biol 135*, 867-881.

*Hum Mol Genet* 4, 951-958.

389-397.

*Neuromuscul Disord* 8, 126-130.

facioscapulohumeral muscular dystrophy. *Am J Hum Genet 81*, 884-894. Lemmers, R.J., van der Vliet, P.J., Klooster, R., Sacconi, S., Camano, P., Dauwerse, J.G.,

*Mol Genet* 7, 1207-1214.

*Neurol* 50, 816-819.

Neurol *55*, 845-850.

1650-1653.

Dauwerse, H.G., Hewitt, J., Hofker, M., Bakker, E., Padberg, G.W., *et al.* (1998). Inter- and intrachromosomal sub-telomeric rearrangements on 4q35: implications for facioscapulohumeral muscular dystrophy (FSHD) aetiology and diagnosis. *Hum* 

Frants, R.R., & van der Maarel, S.M. (2001). Complete allele information in the diagnosis of facioscapulohumeral muscular dystrophy by triple DNA analysis. *Ann* 

Maarel, S.M. (2004). Somatic mosaicism in FSHD often goes undetected. Ann

de Knijff, P., Padberg, G.W., Frants, R.R., & van der Maarel, S.M. (2007). Specific sequence variations within the 4q35 region are associated with

Snider, L., Straasheijm, K.R., van Ommen, G.J., Padberg, G.W.*, et al.* (2010a). A unifying genetic model for facioscapulohumeral muscular dystrophy. *Science 329*,

van der Maarel, S.M. (2010b). Worldwide population analysis of the 4q and 10q subtelomeres identifies only four discrete interchromosomal sequence transfers in

analysis of the mouse adenine nucleotide translocase 1 and 2 genes. *Gene 254*, 57-66.

muscular dystrophy region gene-1 (FRG-1) is an actin-bundling protein associated

Structure, subnuclear distribution, and nuclear matrix association of the

Upadhyaya, M. (1995). Correlation between fragment size at D4F104S1 and age at onset or at wheelchair use, with a possible generational effect, accounts for much phenotypic variation in 4q35-facioscapulohumeral muscular dystrophy (FSHD).

Dystrophy: Molecular Studies 19-21 July 1996, Naarden, The Netherlands.

is a member of a dispersed family of homeobox-containing repeats, subsets of which are clustered on the short arms of the acrocentric chromosomes. *Genomics* 28,


Kohler, J., Rupilius, B., Otto, M., Bathke, K., & Koch, M.C. (1996). Germline mosaicism in

Kornberg, R.D. (1974) Chromatin structure: a repeating unit of histones and DNA. *Science*

Kowaljow, V., Marcowycz, A., Ansseau, E., Conde, C.B., Sauvage, S., Matteotti, C., Arias, C.,

Klooster, R., Straasheijm, K., Shah, B., Sowden, J., Frants, R., Thornton, C., Tawil, R., & van

Klingenberg, M., & Aquila, H. (1982). Some characteristics of the isolated ADP/ATP carrier.

Kondo, T., Bobek, M.P., Kuick, R., Lamb, B., Zhu, X., Narayan, A., Bourc'his, D., Viegas-

Krasnianski, M., Eger, K., Neudecker, S., Jakubiczka, S., & Zierz, S. (2003). Atypical

Lamperti, C., Fabbri, G., Vercelli, L., D'Amico, R., Frusciante, R., Bonifazi, E., Fiorillo, C.,

Laoudj-Chenivesse, D., Carnac, G., Bisbal, C., Hugon, G., Bouillot, S., Desnuelle, C.,

Lehming, N., Le Saux, A., Schuller, J., & Ptashne, M. (1998). Chromatin components as part

Lemmers, R.J., de Kievit, P., Sandkuijl, L., Padberg, G.W., van Ommen, G.J., Frants, R.R., &

Lemmers, R.J., Osborn, M., Haaf, T., Rogers, M., Frants, R.R., Padberg, G.W., Cooper, D.N.,

gene region on chromosome 4q35. *Muscle Nerve* 2, S6-13.

locus encodes a pro-apoptotic protein. *Neuromuscul Disord* 17, 611-623. Kim, J., Kollhoff, A., Bergmann, A., & Stubbs, L. (2003). Methylation-sensitive binding of

at the mRNA and protein level. *Eur J Hum Genet 17*, 1615-1624.

predominantly in oogenesis. *Hum Genet* 98, 485-490.

imprinted gene, Peg3. *Hum Mol Genet* 12, 233-245.

Tokai J Exp Clin Med *7 Suppl*, 43-49.

deletion. *Arch Neurol* 60, 1421-1425.

score. *Muscle Nerve* 42, 213-217.

7326.

236.

*Neurology* 61, 178-183.

*Hum Mol Genet* 9, 597-604.

184, 868-871.

4q35 facioscapulohumeral muscular dystrophy (FSHD1A) occurring

Corona, E.D., Nunez, N.G., Leo, O.*, et al.* (2007). The DUX4 gene at the FSHD1A

transcription factor YY1 to an insulator sequence within the paternally expressed

der Maarel, S. (2009). Comprehensive expression analysis of FSHD candidate genes

Pequignot, E., Ehrlich, M., & Hanash, S.M. (2000). Whole-genome methylation scan in ICF syndrome: hypomethylation of non-satellite DNA repeats D4Z4 and NBL2.

phenotypes in patients with facioscapulohumeral muscular dystrophy 4q35

Borsato, C., Cao, M., Servida, M., *et al.* (2010). A standardized clinical evaluation of patients affected by facioscapulohumeral muscular dystrophy: The FSHD clinical

Vassetzky, Y., & Fernandez, A. (2005). Increased levels of adenine nucleotide translocator 1 protein and response to oxidative stress are early events in facioscapulohumeral muscular dystrophy muscle. *J Mol Med* (Berl) 83, 216-224. Lee, J.H., Goto, K., Matsuda, C., & Arahata, K. (1995). Characterization of a tandemly

repeated 3.3-kb KpnI unit in the facioscapulohumeral muscular dystrophy (FSHD)

of a putative transcriptional repressing complex. *Proc Natl Acad Sci U S A* 95, 7322-

van der Maarel, S.M. (2002). Facioscapulohumeral muscular dystrophy is uniquely associated with one of the two variants of the 4q subtelomere. Nat Genet *32*, 235-

van der Maarel, S.M., & Upadhyaya, M. (2003). D4F104S1 deletion in facioscapulohumeral muscular dystrophy: phenotype, size, and detection.


Facioscapulohumeral Muscular Dystrophy: From Clinical Data to Molecular Genetics and Return 49

Peterson, C.L., & Laniel, M.A. (2004). Histones and histone modifications. *Curr Biol* 14,

Petrov, A., Allinne, J., Pirozhkova, I., Laoudj, D., Lipinski, M., & Vassetzky, Y.S. (2008). A

Petrov, A., Pirozhkova, I., Carnac, G., Laoudj, D., Lipinski, M., & Vassetzky, Y.S. (2006).

Pirozhkova, I., Petrov, A., Dmitriev, P., Laoudj, D., Lipinski, M., & Vassetzky, Y. (2008). A

Rappsilber, J., Ryder, U., Lamond, A.I., & Mann, M. (2002). Large-scale proteomic analysis of

Ricci, E., Galluzzi, G., Deidda, G., Cacurri, S., Colantoni, L., Merico, B., Piazzo, N., Servidei,

Robertson, K.D., & Wolffe, A.P. (2000). DNA methylation in health and disease. *Nat Rev* 

Rossi, M., Ricci, E., Colantoni, L., Galluzzi, G., Frusciante, R., Tonali, P.A., & Felicetti, L.

Rudnik-Schoneborn, S., Weis, J., Kress, W., Hausler, M., & Zerres, K. (2008). Becker's

Saenz, A., Leturcq, F., Cobo, A.M., Poza, J.J., Ferrer, X., Otaegui, D., Camano, P., Urtasun,

Scionti, I., Fabbri, G., Fiorillo, C., Ricci, G., Greco, F., D'Amico, R., Tremanini, A., Vercelli, L.,

the human spliceosome. *Genome Res* 12, 1231-1245.

*Neuromuscul Disord* 18, 881-885.

pressure. BMC Med Genet *8*, 8.

881-885.

732-742.

myoblast cultures of FSHD patients. *J Med Genet* 41, 826-836.

nuclear matrix attachment site in the 4q35 locus has an enhancer-blocking activity in vivo: implications for the facio-scapulo-humeral dystrophy. *Genome Res 18*,

Chromatin loop domain organization within the 4q35 locus in facioscapulohumeral dystrophy patients versus normal human myoblasts. *Proc Natl Acad Sci U S A* 103,

functional role for 4qA/B in the structural rearrangement of the 4q35 region and in the regulation of FRG1 and ANT1 in facioscapulohumeral dystrophy. *PLoS One* 3,

S., Vigneti, E., Pasceri, V., *et al.* (1999). Progress in the molecular diagnosis of facioscapulohumeral muscular dystrophy and correlation between the number of KpnI repeats at the 4q35 locus and clinical phenotype. *Ann Neurol* 45, 751-757. Rijkers, T., Deidda, G., van Koningsbruggen, S., van Geel, M., Lemmers, R.J., van Deutekom,

J.C., Figlewicz, D., Hewitt, J.E., Padberg, G.W., Frants, R.R., *et al.* (2004). FRG2, an FSHD candidate gene, is transcriptionally upregulated in differentiating primary

*Genet 1,* 11-19.Rudnik-Schoneborn, S., Weis, J., Kress, W., Hausler, M., and Zerres, K. (2008). Becker's muscular dystrophy aggravating facioscapulohumeral muscular dystrophy--double trouble as an explanation for an atypical phenotype.

(2007). The Facioscapulohumeral muscular dystrophy region on 4qter and the homologous locus on 10qter evolved independently under different evolutionary

muscular dystrophy aggravating facioscapulohumeral muscular dystrophy- double trouble as an explanation for an atypical phenotype. *Neuromuscul Disord* 18,

M., Vilchez, J., Gutierrez-Rivas, E., *et al.* (2005). LGMD2A: genotype-phenotype correlations based on a large mutational survey on the calpain 3 gene. *Brain* 128,

Tomelleri, G., Cao, M., Santoro, L., Percesepe, A., & Tupler, R. (2012a). Facioscapulohumeral muscular dystrophy: new insights from compound heterozygotes and implication for prenatal genetic counselling.*JMG* january 2012

R546-551.

39-45.

6982-6987.

e3389.


Mal, A.K. (2006). Histone methyltransferase Suv39h1 represses MyoD-stimulated myogenic

Masny, P.S., Bengtsson, U., Chung, S.A., Martin, J.H., van Engelen, B., van der Maarel, S.M.,

Masny, P.S., Chan, O.Y., de Greef, J.C., Bengtsson, U., Ehrlich, M., Tawil, R., Lock, L.F.,

Matsumura, T., Goto, K., Yamanaka, G., Lee, J.H., Zhang, C., Hayashi, Y.K., & Arahata, K.

Mostacciuolo, M.L., Pastorello, E., Vazza, G., Miorin, M., Angelini, C., Tomelleri, G.,

Nadaj-Pakleza, A.A., Vincitorio, C.M., Laforet, P., Eymard, B., Dion, E., Teijeira, S., Vietez, I.,

Nagele, R.G., Velasco, A.Q., Anderson, W.J., McMahon, D.J., Thomson, Z., Fazekas, J., Wind,

Osborne, R.J., Welle, S., Venance, S.L., Thornton, C.A., & Tawil, R. (2007). Expression profile

Ottaviani, A., Schluth-Bolard, C., Gilson, E., & Magdinier, F. (2010). D4Z4 as a prototype of CTCF and lamins-dependent insulator in human cells. *Nucleus* 1, 30-36. Ottaviani, A., Schluth-Bolard, C., Rival-Gervier, S., Boussouar, A., Rondier, D., Foerster,

Oya, Y., Morita, H., Ogawa, M., Nonaka, I., Tsujino, S., & Kawai, M. (2001). [Adult form of

Padberg, G.W., Frants, R.R., Brouwer, O.F., Wijmenga, C., Bakker, E., & Sandkuijl, L.A.

Padberg, G.W., Lunt, P.W., Koch, M., & Fardeau, M. (1991). Diagnostic criteria for facioscapulohumeral muscular dystrophy. *Neuromuscul Disord* 1, 231-234.

requires A-type lamins and CTCF. *EMBO J* 28, 2428-2436.

Neurol Neurosurg 94 Suppl, S21-24.

facioscapulohumeral dystrophy]. *Rinsho Shinkeigaku* 41, 390-396. Padberg, G. (1982). Facioscapulohumeral disease. Leiden University The Netherlands. Padberg, G.W. (1992). Why cells die in facioscapulohumeral muscular dystrophy. Clin

maintenance of interphase chromosome topology. *J Cell Sci 114*, 377-388. Nightingale, K.P., O'Neill, L.P., & Turner, B.M. (2006). Histone modifications: signalling

nuclear envelope disease? *Hum Mol Genet* 13, 1857-1871.

populations and FSHD patients. *BMC Neurol* 2, 7.

McArdle disease. *Muscle Nerve* 40, 350-357.

*Clin Genet*. 2009 Jun;75(6):550-5

& Winokur, S.T. (2004). Localization of 4q35.2 to the nuclear periphery: is FSHD a

Hewitt, J.E., Stocksdale, J., Martin, J.H., *et al.* (2010). Analysis of allele-specific RNA transcription in FSHD by RNA-DNA FISH in single myonuclei. *Eur J Hum Genet* 18,

(2002). Chromosome 4q;10q translocations; comparison with different ethnic

Galluzzi, G. & Trevisan, C.P. (2009). Facioscapulohumeral muscular dystrophy: epidemiological and molecular study in a north-east Italian population sample.

Jeanpierre, M., Navarro, C., & Stojkovic, T. (2009). Permanent muscle weakness in

K., & Lee, H. (2001). Telomere associations in interphase nuclei: possible role in

receptors and potential elements of a heritable epigenetic code. *Curr Opin Genet Dev*

of FSHD supports a link between retinal vasculopathy and muscular dystrophy.

A.M., Morere, J., Bauwens, S., Gazzo, S., Callet-Bauchu, E., *et al.* (2009). Identification of a perinuclear positioning element in human subtelomeres that

acid maltase deficiency presenting with pattern of muscle weakness resembling

(1995). Facioscapulohumeral muscular dystrophy in the Dutch population. *Muscle* 

differentiation. *EMBO J* 25, 3323-3334.

448-456.

16, 125-136.

*Neurology* 68, 569-577.

*Nerve* 2, S81-84.


Facioscapulohumeral Muscular Dystrophy: From Clinical Data to Molecular Genetics and Return 51

Tonini, M.M., Passos-Bueno, M.R., Cerqueira, A., Matioli, S.R., Pavanello, R., & Zatz, M.

Trevisan, C.P., Pastorello, E., Tomelleri, G., Vercelli, L., Bruno, C., Scapolan, S., Siciliano, G.,

Tsien, F., Sun, B., Hopkins, N.E., Vedanarayanan, V., Figlewicz, D., Winokur, S., & Ehrlich,

Tupler, R., Barbierato, L., Memmi, M., Sewry, C.A., De Grandis, D., Maraschio, P., Tiepolo,

Tupler, R., Berardinelli, A., Barbierato, L., Frants, R., Hewitt, J.E., Lanzi, G., Maraschio, P., &

Upadhyaya, M., Maynard, J., Osborn, M., Jardine, P., Harper, P.S., & Lunt, P. (1995).

van der Maarel, S.M., Deidda, G., Lemmers, R.J., Bakker, E., van der Wielen, M.J., Sandkuijl,

facioscapulohumeral muscular dystrophy (FSHD). *J Med Genet* 36, 823-828. van der Maarel, S.M., Deidda, G., Lemmers, R.J., van Overveld, P.G., van der Wielen, M.,

interaction between chromosomes 4 and 10. *Am J Hum Genet* 66, 26-35. van der Maarel, S.M., Frants, R.R., & Padberg, G.W. (2007). Facioscapulohumeral muscular

van Deutekom, J.C., Wijmenga, C., van Tienhoven, E.A., Gruter, A.M., Hewitt, J.E., Padberg,

van Deutekom, J.C.(1996a) Toward the molecular mechanism of faciuscapulohumeral

muscular dystrophy. *PhD Thesis*,Leiden University, Leiden

(FSHD) with different clinical expression. *J Med Genet* 35, 778-783.

normal and FSHD cell cultures and tissues. *Mol Genet Metab* 74, 322-331. Tsuji, M., Kinoshita, M., Imai, Y., Kawamoto, M., & Kohara, N. (2009). Facioscapulohumeral

muscular dystrophy (FSHD). *Neuromuscul Disord* 14, 33-38.

1453-1465.

1353-1358.

*Nerve* 2, S45-49.

*Neuromuscul Disord* 19, 140-142.

muscular dystrophy. *J Med Genet* 33, 366-370.

dystrophy. *Biochim Biophys Acta* 1772, 186-194.

repeated unit. *Hum Mol Genet 2*, 2037-2042.

dystrophy (FSHD) disease expression is almost exclusively associated with an FSHD locus located on a 4qA-defined 4qter subtelomere. J Med Genet *44*, 215-218. Tolhuis, B., Palstra, R.J., Splinter, E., Grosveld, F., & de Laat, W. (2002). Looping and

interaction between hypersensitive sites in the active beta-globin locus. *Mol Cell* 10,

(2004). Asymptomatic carriers and gender differences in facioscapulohumeral

& Comacchio, F. (2008). Facioscapulohumeral muscular dystrophy: hearing loss and other atypical features of patients with large 4q35 deletions. *Eur J Neurol 1*5,

M. (2001). Methylation of the FSHD syndrome-linked subtelomeric repeat in

muscular dystrophy presenting with hypertrophic cardiomyopathy: a case study.

L., & Ferlini, A. (1998). Identical de novo mutation at the D4F104S1 locus in monozygotic male twins affected by facioscapulohumeral muscular dystrophy

Tiepolo, L. (1996). Monosomy of distal 4q does not cause facioscapulohumeral

Germinal mosaicism in facioscapulohumeral muscular dystrophy (FSHD). *Muscle* 

L., Hewitt, J.E., Padberg, G.W., & Frants, R.R. (1999). A new dosage test for subtelomeric 4;10 translocations improves conventional diagnosis of

Hewitt, J.E., Sandkuijl, L., Bakker, B., van Ommen, G.J., Padberg, G.W., *et al.* (2000). De novo facioscapulohumeral muscular dystrophy: frequent somatic mosaicism, sex-dependent phenotype, and the role of mitotic transchromosomal repeat

G.W., van Ommen, G.J., Hofker, M.H., & Frants, R.R. (1993). FSHD associated DNA rearrangements are due to deletions of integral copies of a 3.2 kb tandemly


Scionti, I., Greco, F., Ricci, G., Govi, M., Arashiro, P., Vercelli, L., Berardinelli, A., Angelini,

the current criteria for the molecular diagnosis of fascioscapulohumeral muscular dystrophy

Sharer, J.D., Shern, J.F., Van Valkenburgh, H., Wallace, D.C., & Kahn, R.A. (2002). ARL2 and

Snider, L., Geng, L.N., Lemmers, R.J., Kyba, M., Ware, C.B., Nelson, A.M., Tawil, R.,

Spilianakis, C.G., Lalioti, M.D., Town, T., Lee, G.R., & Flavell, R.A. (2005). Interchromosomal associations between alternatively expressed loci. *Nature* 435, 637-645. Stepien, G., Torroni, A., Chung, A.B., Hodge, J.A., & Wallace, D.C. (1992). Differential

Strahl, B.D., & Allis, C.D. (2000). The language of covalent histone modifications. *Nature* 403,

Sun, C.Y., van Koningsbruggen, S., Long, S.W., Straasheijm, K., Klooster, R., Jones, T.I.,

Sun, H.B., Shen, J., & Yokota, H. (2000). Size-dependent positioning of human chromosomes

Swinkels, B.W., Gould, S.J., & Subramani, S. (1992). Targeting efficiencies of various

Tam, R., Smith, K.P., & Lawrence, J.B. (2004). The 4q subtelomere harboring the FSHD locus

Tanabe, H., Habermann, F.A., Solovei, I., Cremer, M., & Cremer, T. (2002). Non-random

Tawil, R., Forrester, J., Griggs, R.C., Mendell, J., Kissel, J., McDermott, M., King, W.,

Tawil, R., van der Maarel, S., Padberg, G.W., & van Engelen, B.G. (2010). 171st ENMC

facioscapulohumeral muscular dystrophy. *Neuromuscul Disord* 20, 471-475. Thomas, N.S., Wiseman, K., Spurlock, G., MacDonald, M., Ustek, D., & Upadhyaya, M.

considerations and functional implications. *Mutat Res 504*, 37-45.

during muscle cell differentiation. *J Biol Chem 267*, 14592-14597.

Associated and Actin-Bundling Protein. *J Mol Biol*.

in interphase nuclei. *Biophys J 79*, 184-190.

*FEBS* Lett 305, 133-136.

telomeres. *J Cell Biol 167*, 269-279.

DY Group. *Ann Neurol* 39, 744-748.

analysis challenges

*Cell 13*, 71-83.

41-45.

(FSHD). *AJHG* Paper accepted.

gene. PLoS Genet *6*, e1001181.

C., Antonini, G., Cao, M., Di Muzio, A., Moggio, M., Morandi, L., Ricci, E., Rodolico, C., Ruggero, L., Santoro, L., Siciliano, G., Tomelleri, G., Trevisan, CP., Galluzzi, G., Wright, W., Zatz, M., & Tupler, R. (2012b). Large scale population

BART enter mitochondria and bind the adenine nucleotide transporter*. Mol Biol* 

Filippova, G.N., van der Maarel, S.M., Tapscott, S.J.*, et al.* (2010). Facioscapulohumeral dystrophy: incomplete suppression of a retrotransposed

expression of adenine nucleotide translocator isoforms in mammalian tissues and

Bellini, M., Levesque, L., Brieher, W.M., van der Maarel, S.M.*, et al.* (2011). Facioscapulohumeral Muscular Dystrophy Region Gene 1 Is a Dynamic RNA-

permutations of the consensus C-terminal tripeptide peroxisomal targeting signal.

is specifically anchored with peripheral heterochromatin unlike most human

radial arrangements of interphase chromosome territories: evolutionary

Weiffenbach, B., & Figlewicz, D. (1996). Evidence for anticipation and association of deletion size with severity in facioscapulohumeral muscular dystrophy. The FSH-

international workshop: Standards of care and management of

(2007). A large patient study confirming that facioscapulohumeral muscular

dystrophy (FSHD) disease expression is almost exclusively associated with an FSHD locus located on a 4qA-defined 4qter subtelomere. J Med Genet *44*, 215-218.


Facioscapulohumeral Muscular Dystrophy: From Clinical Data to Molecular Genetics and Return 53

Wijmenga, C., Frants, R.R., Brouwer, O.F., Moerer, P., Weber, J.L., & Padberg, G.W. (1990).

Wijmenga, C., Brouwer, O.F., Padberg, G.W., & Frants, R.R. (1992a). Transmission of de-

Wijmenga, C., Hewitt, J.E., Sandkuijl, L.A., Clark, L.N., Wright, T.J., Dauwerse, H.G., Gruter,

Winokur, S.T., Bengtsson, U., Feddersen, J., Mathews, K.D., Weiffenbach, B., Bailey, H.,

Winokur, S.T., Barrett, K., Martin, J.H., Forrester, J.R., Simon, M., Tawil, R., Chung, S.A.,

Winokur, S.T., Chen, Y.W., Masny, P.S., Martin, J.H., Ehmsen, J.T., Tapscott, S.J., van der

Wohlgemuth, M., Lemmers, R.J., van der Kooi, E.L., van der Wielen, M.J., van Overveld,

Xu, G.L., Bestor, T.H., Bourc'his, D., Hsieh, C.L., Tommerup, N., Bugge, M., Hulten, M., Qu,

Yang, F., Shao, C., Vedanarayanan, V., & Ehrlich, M. (2004). Cytogenetic and immuno-FISH

Zatz, M., Marie, S.K., Cerqueira, A., Vainzof, M., Pavanello, R.C., & Passos-Bueno, M.R.

facioscapulohumeral muscular dystrophy. *Chromosoma* 112, 350-359. Yao, Y.L., Yang, W.M., & Seto, E. (2001). Regulation of transcription factor YY1 by

acetylation and deacetylation. *Mol Cell Biol* 21, 5979-5991.

*Lancet* 336, 651-653.

340, 985-986.

Genet 2, 26-30.

225-234.

*Neuromuscul Disord* 13, 322-333.

FSHD-sized 4q35 alleles. *Neurology* 61, 909-913.

*Genet* 12, 2895-2907.

gene. *Nature* 402, 187-191.

families. *Am J Hum Genet* 56, 99-105.

Location of facioscapulohumeral muscular dystrophy gene on chromosome 4.

novo mutation associated with facioscapulohumeral muscular dystrophy. *Lancet*

A.M., Hofker, M.H., Moerer, P., Williamson, R., *et al.* (1992b). *Chromosome* 4q DNA rearrangements associated with facioscapulohumeral muscular dystrophy. Nat

Markovich, R.P., Murray, J.C., Wasmuth, J.J., Altherr, M.R., *et al.* (1994). The DNA rearrangement associated with facioscapulohumeral muscular dystrophy involves a heterochromatin-associated repetitive element: implications for a role of chromatin structure in the pathogenesis of the disease. *Chromosome* Res 2,

Masny, P.S., & Figlewicz, D.A. (2003a). Facioscapulohumeral muscular dystrophy (FSHD) myoblasts demonstrate increased susceptibility to oxidative stress.

Maarel, S.M., Hayashi, Y., & Flanigan, K.M. (2003b). Expression profiling of FSHD muscle supports a defect in specific stages of myogenic differentiation. *Hum Mol* 

P.G., Dauwerse, H., Bakker, E., Frants, R.R., Padberg, G.W., & van der Maarel, S.M. (2003). Possible phenotypic dosage effect in patients compound heterozygous for

X., Russo, J.J., & Viegas-Pequignot, E. (1999). Chromosome instability and immunodeficiency syndrome caused by mutations in a DNA methyltransferase

analysis of the 4q subtelomeric region, which is associated with

(1998). The facioscapulohumeral muscular dystrophy (FSHD1) gene affects males more severely and more frequently than females. *Am J Med Genet* 77, 155-161. Zatz, M., Marie, S.K., Passos-Bueno, M.R., Vainzof, M., Campiotto, S., Cerqueira, A.,

Wijmenga, C., Padberg, G., & Frants, R. (1995). High proportion of new mutations and possible anticipation in Brazilian facioscapulohumeral muscular dystrophy


van Deutekom, J.C., Bakker, E., Lemmers, R.J., van der Wielen, M.J., Bik, E., Hofker, M.H.,

genetic counselling and etiology of FSHD1. *Hum Mol Genet* 5, 1997-2003. van Deutekom, J.C., Lemmers, R.J., Grewal, P.K., van Geel, M., Romberg, S., Dauwerse,

van Geel, M., Heather, L.J., Lyle, R., Hewitt, J.E., Frants, R.R., & de Jong, P.J. (1999). The

van Koningsbruggen, S., Dirks, R.W., Mommaas, A.M., Onderwater, J.J., Deidda, G.,

van Overveld, P.G., Lemmers, R.J., Deidda, G., Sandkuijl, L., Padberg, G.W., Frants, R.R., &

van Overveld, P.G., Lemmers, R.J., Sandkuijl, L.A., Enthoven, L., Winokur, S.T., Bakels, F.,

van Overveld, P.G., Enthoven, L., Ricci, E., Rossi, M., Felicetti, L., Jeanpierre, M., Winokur,

Vitelli, F., Villanova, M., Malandrini, A., Bruttini, M., Piccini, M., Merlini, L., Guazzi, G., &

Weierich, C., Brero, A., Stein, S., von Hase, J., Cremer, C., Cremer, T., & Solovei, I. (2003).

Weiffenbach, B., Dubois, J., Storvick, D., Tawil, R., Jacobsen, S.J., Gilbert, J., Wijmenga, C.,

Weiffenbach, B., bagley,R., Falls, K., Hyser, C. & Storvick, D., (1992). Linkage analyses of

and murine lymphocytes*. Chromosome Res 11*, 485-502.

(FSHD) gene to distal 4q35. *Am J Hum Genet* 51, 416-423.

recombination events. *Nat Genet* 4, 165-169.

facioscapulohumeral muscular dystrophy. *Muscle Nerve* 22, 1437-1441. Wallace, L.M., Garwick, S.E., Mei, W., Belayew, A., Coppee, F., Ladner, K.J., Guttridge, D.,

4q telomeres suggests a common origin. *Genomics* 79, 210-217.

the nucleolus, Cajal bodies, and speckles. *J Med Genet* 41, e46.

chromosome 4q35. *Hum Mol Genet 5*, 581-590.

muscular dystrophy. *Nat Genet* 35, 315-317.

2879-2884.

*58*, 569-576.

540-552.

Padberg, G.W., & Frants, R.R. (1996b). Evidence for subtelomeric exchange of 3.3 kb tandemly repeated units between chromosomes 4q35 and 10q26: implications for

H.G., Wright, T.J., Padberg, G.W., Hofker, M.H., Hewitt, J.E.*, et al.* (1996c). Identification of the first gene (FRG1) from the FSHD region on human

FSHD region on human chromosome 4q35 contains potential coding regions among pseudogenes and a high density of repeat elements. *Genomics* 61, 55-65. van Geel, M., Dickson, M.C., Beck, A.F., Bolland, D.J., Frants, R.R., van der Maarel, S.M., de

Jong, P.J., & Hewitt, J.E. (2002). Genomic analysis of human chromosome 10q and

Padberg, G.W., Frants, R.R., & van der Maarel, S.M. (2004). FRG1P is localised in

van der Maarel, S.M. (2000). Interchromosomal repeat array interactions between chromosomes 4 and 10: a model for subtelomeric plasticity. *Hum Mol Genet* 9,

Padberg, G.W., van Ommen, G.J., Frants, R.R., & van der Maarel, S.M. (2003). Hypomethylation of D4Z4 in 4q-linked and non-4q-linked facioscapulohumeral

S.T., Frants, R.R., Padberg, G.W., & van der Maarel, S.M. (2005). Variable hypomethylation of D4Z4 in facioscapulohumeral muscular dystrophy. *Ann Neurol* 

Renieri, A. (1999). Inheritance of a 38-kb fragment in apparently sporadic

Yang, J., & Harper, S.Q. (2011). DUX4, a candidate gene for facioscapulohumeral muscular dystrophy, causes p53-dependent myopathy in vivo. *Ann Neurol 69*,

Three-dimensional arrangements of centromeres and telomeres in nuclei of human

Mendell, J.R., Winokur, S., Altherr, M.R., *et al.* (1993). Mapping the facioscapulohumeral muscular dystrophy gene is complicated by chromsome 4q35

five chromosome 4 markers localizes the facioscapulohumeral muscular dystrophy


**3** 

**AON-Mediated Exon Skipping** 

*Department of Human Genetics, Leiden University Medical Center* 

*The Netherlands* 

**for Duchenne Muscular Dystrophy** 

Ingrid E. C. Verhaart and Annemieke Aartsma-Rus

Duchenne muscular dystrophy (DMD) is a genetic, X-chromosome recessive, severe and progressive muscle wasting disorder, affecting around 1 in 3500 newborn boys. The onset of the disease is in early childhood and, nowadays, most children are diagnosed before the age of 5. The first signs of muscular weakness become apparent around the age of 2 or 3 years. In most patients the age at which the child starts to walk is delayed (retarded motor development). The children have less endurance and difficulties with running and climbing stairs (Moser, 1984). Gower's sign is a reflection of the weakness of the muscles of the lower extremities (knee and hip extensors): the child helps himself to get upright from sitting position by using his upper extremities: first by rising to stand on his arms and knees, and then "walking" his hands up his legs to stand upright (Gowers, 1895). Muscle wasting is often symmetrical, however not all muscles are affected to the same extent. A prominent feature of the disease is enlargement of the calve muscle, caused by replacement of muscle fibres by connective and adipose tissue. Furthermore, the pelvic girdle, trunk and abdomen are severely affected and to a lesser extent the shoulder girdle and proximal muscles of the upper extremities. Progressive weakness and contractures of the leg muscles lead to wheelchair-dependency around the age of 10. Thereafter the muscle contractions increase rapidly leading to spinal deformities and scoliosis, often with an asymmetric distribution pattern. Involvement of the intercostal muscles and distortion of the thorax lead to respiratory failure and patients often require assisted ventilation in the mid to late teens. Thereafter dilated cardiomyopathy becomes apparent and most patients die before the age of 30. Another common feature is mental retardation (IQ less than 70) in around 20-30% of

Becker Muscular Dystrophy (BMD) is a related, but much milder, form of muscular weakness, affecting around 1 in 20 000 men. The phenotype varies between individual patients, from very mild to moderately severe, but the course of the disease is more benign compared to DMD. On average, the age of onset is around 12 years; however some patients remain asymptomatic until much higher ages. The age of wheelchair-dependency also shows more variability, but in general is in their second or third decade of life. The most severely affected patients die between 40 and 50 years of age, whereas patients with a mild

**1. Introduction** 

the patients (Emery, 2002).

