Fascioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant neuromuscular disorder linked to partial deletion of integral numbers of a 3.3 kb polymorphic repeat, D4Z4, within the subtelomeric region of chromosome 4q. Although the relationship between deletions of D4Z4 and FSHD is well established, how this triggers the disease remains unclear. We have mapped the DNA loop domain containing the D4Z4 repeat cluster in human primary myoblasts and in murine-human hybrids. A nuclear matrix attachment site was found located in the vicinity of the repeat. Prominent in normal human myoblasts and nonmuscular human cells, this site is much weaker in muscle cells derived from FSHD patients, suggesting that the D4Z4 repeat array and upstream genes reside in two loops in nonmuscular cells and normal human myoblasts but in only one loop in FSHD myoblasts. We propose a model whereby the nuclear scaffold͞matrix attached region regulates chromatin accessibility and expression of genes implicated in the genesis of FSHD. The disorder is related to a short repeat array that remains after deletion of an integral number of tandemly arrayed 3.3-kb repeat units on chromosome 4. The size of this polymorphic locus (D4Z4) varies in normal individuals from 35 to 300 kb, whereas in FSHD patients it is consistently shorter than 35 kb (3). Partial deletion of the D4Z4 array on chromosome 4 ultimately leads to FSHD and is currently used as a diagnostic tool in genetic counseling to predict the probability of the disease (1,(3)(4)(5). A correlation exists between the extent of the deletion and its clinical expression: Indeed, patients with one to three repeats develop an early FSHD, whereas individuals with nine to 10 repeats exhibit a weaker form of the disease (5).Extensive efforts to identify gene transcripts associated with the 4q35-specific D4Z4 repeat, as potential FSHD candidate genes, have been largely unsuccessful (6). The 3.3-kb D4Z4 elements contain a cryptic DUX4 gene potentially coding for a double homeodomain protein (7), and an overall perturbation of mRNA expression profiles can be observed in FSHD patients (8-10), but the disease appears to result from an as yet unexplained mechanism with a genetic alteration not residing within a causative gene for the disease.The 4q35 genomic region ( Fig. 1) displays heterochromatic features and might exert repressive effects on neighboring genes with a mechanism similar to position effect variegation. A decreased D4Z4 repeat number consistently results in inappropriate up-regulation of adjacent FRG2, FRG1, and Ant1 in FSHD muscle (11-13). It has also been shown that a transcriptional repressor complex binds D4Z4, whose deletion would trigger overexpression by lack of repression (11). Indeed, overexpression of FRG1 in transgenic mice provokes a phenotype similar to that of FSHD (14). However, such a model of position effect has been recently challenged in two reports of an apparent lack of up-regulation of any 4q35 gene and because of the histone H4 acetylation state in FSHD lymph...

Facio-scapulo-humeral dystrophy (FSHD), a muscular hereditary disease with a prevalence of 1 in 20,000, is caused by a partial deletion of a subtelomeric repeat array on chromosome 4q. Earlier, we demonstrated the existence in the vicinity of the D4Z4 repeat of a nuclear matrix attachment site, FR-MAR, efficient in normal human myoblasts and nonmuscular human cells but much weaker in muscle cells from FSHD patients. We now report that the D4Z4 repeat contains an exceptionally strong transcriptional enhancer at its 5′-end. This enhancer up-regulates transcription from the promoter of the neighboring FRG1 gene. However, an enhancer blocking activity was found present in FR-MAR that in vitro could protect transcription from the enhancer activity of the D4Z4 array. In vivo, transcription from the FRG1 and FRG2 genes could be down- or up-regulated depending on whether or not FR-MAR is associated with the nuclear matrix. We propose a model for an etiological role of the delocalization of FR-MAR in the genesis of FSHD.

The number of D4Z4 repeats in the subtelomeric region of chromosome 4q is strongly reduced in patients with Facio-Scapulo-Humeral Dystrophy (FSHD). We performed chromosome conformation capture (3C) analysis to document the interactions taking place among different 4q35 markers. We found that the reduced number of D4Z4 repeats in FSHD myoblasts was associated with a global alteration of the three-dimensional structure of the 4q35 region. Indeed, differently from normal myoblasts, the 4qA/B marker interacted directly with the promoters of the FRG1 and ANT1 genes in FSHD cells. Along with the presence of a newly identified transcriptional enhancer within the 4qA allele, our demonstration of an interaction occurring between chromosomal segments located megabases away on the same chromosome 4q allows to revisit the possible mechanisms leading to FSHD.

BackgroundIt becomes increasingly evident that nuclesomes are far from being identical to each other. This nucleosome diversity is due partially to the existence of histone variants encoded by separate genes. Among the known histone variants the less characterized are H2A.Bbd and different forms of macroH2A. This is especially true in the case of H2A.Bbd as there are still no commercially available antibodies specific to H2A.Bbd that can be used for chromatin immunoprecipitation (ChIP).MethodsWe have generated HeLa S3 cell lines stably expressing epitope-tagged versions of macroH2A1.1, H2A.Bbd or canonical H2A and analyzed genomic distribution of the tagged histones using ChIP-on-chip technique.ResultsThe presence of histone H2A variants macroH2A1.1 and H2A.Bbd has been analyzed in the chromatin of several segments of human chromosomes 11, 16 and X that have been chosen for their different gene densities and chromatin status. Chromatin immunoprecipitation (ChIP) followed by hybridization with custom NimbleGene genomic microarrays demonstrated that in open chromatin domains containing tissue-specific along with housekeeping genes, the H2A.Bbd variant was preferentially associated with the body of a subset of transcribed genes. The macroH2A1.1 variant was virtually absent from some genes and underrepresented in others. In contrast, in closed chromatin domains which contain only tissue-specific genes inactive in HeLa S3 cells, both macroH2A1.1 and H2A.Bbd histone variants were present and often colocalized.ConclusionsGenomic distribution of macro H2A and H2A.Bbd does not follow any simple rule and is drastically different in open and closed genomic domains.

Satellite DNA, also known as tandemly repeated DNA, consists of clusters of repeated sequences and represents a diverse class of highly repetitive elements. Satellite DNA can be divided into several classes according to the size of an individual repeat: microsatellites, minisatellites, midisatellites, and macrosatellites. Originally considered as «junk» DNA, satellite DNA has more recently been reconsidered as having various functions. Moreover, due to the repetitive nature of the composing elements, their presence in the genome is associated with high frequency mutations, epigenetic changes and modifications in gene expression patterns, with a potential to lead to human disease. Therefore, the satellite DNA study will be beneficial for developing a treatment of satelliterelated diseases, such as FSHD, neurological, developmental disorders and cancers.

Facioscapulohumeral dystrophy (FSHD) is an epi/genetic satellite disease associated with at least two satellite sequences in 4q35: (i) D4Z4 macrosatellite and (ii) β-satellite repeats (BSR), a prevalent part of the 4qA allele. Most of the recent FSHD studies have been focused on a DUX4 transcript inside D4Z4 and its tandem contraction in FSHD patients. However, the D4Z4-contraction alone is not pathological, which would also require the 4qA allele. Since little is known about BSR, we investigated the 4qA BSR functional role in the transcriptional control of the FSHD region 4q35. We have shown that an individual BSR possesses enhancer activity leading to activation of the Adenine Nucleotide Translocator 1 gene (ANT1), a major FSHD candidate gene. We have identified ZNF555, a previously uncharacterized protein, as a putative transcriptional factor highly expressed in human primary myoblasts that interacts with the BSR enhancer site and impacts the ANT1 promoter activity in FSHD myoblasts. The discovery of the functional role of the 4qA allele and ZNF555 in the transcriptional control of ANT1 advances our understanding of FSHD pathogenesis and provides potential therapeutic targets.

No abstract

No abstract