Sequence Homology between 4qter and 10qter Loci Facilitates the Instability of Subtelomeric KpnI Repeat Units Implicated in Facioscapulohumeral Muscular Dystrophy

Abstract

Physical mapping and in situ hybridization experiments have shown that a duplicated locus with a structural organization similar to that of the 4q35 locus implicated in facioscapulohumeral muscular dystrophy is present in the subtelomeric portion of 10q. We performed sequence analysis of the p13E-11 probe and of the adjacent KpnI tandem-repeat unit derived from a 10qter cosmid clone and compared our results with those published, by other laboratories, for the 4q35 region. We found that the sequence homology range is 98%-100% and confirmed that the only difference that can be exploited for differentiation of the 10qter from the 4q35 alleles is the presence of an additional BlnI site within the 10qter KpnI repeat unit. In addition, we observed that the high degree of sequence homology does facilitate interchromosomal exchanges resulting in displacement of the whole set of BlnI-resistant or BlnI-sensitive KpnI repeats from one chromosome to the other. However, partial translocations escape detection if the latter simply relies on the hybridization pattern from double digestion with EcoRI/BlnI and with p13E-11 as a probe. We discovered that the restriction enzyme Tru9I cuts at both ends of the array of KpnI repeats of different chromosomal origins and allows the use of cloned KpnI sequences as a probe by eliminating other spurious fragments. This approach coupled with BlnI digestion permitted us to investigate the structural organization of BlnI-resistant and BlnI-sensitive units within translocated chromosomes of 4q35 and 10q26 origin. A priori, the possibility that partial translocations could play a role in the molecular mechanism of the disease cannot be excluded.

Molecular basis of facioscapulohumeral muscular dystrophy

“…The reverse configuration of chromosome 4-type repeats on chromosome 10 is equally frequent [34,35], and hybrid repeats, consisting of both 4-type and 10-type repeat units, have also been identified. These data demonstrate that recombination between 4q35 and 10q26 occurs frequently, and show the dynamic characteristics of the 4q-10q homologous domain [36,37]. Most important, these observations confirm that FSHD is not caused by alteration of the specific sequence composition of the D4Z4 repeats but rather by their structural organization in the 4q35 region.…”

Subtelomeric DNA hypomethylation is not required for telomeric sister chromatid exchanges in ALT cells

“…The ALT phenotype of LB188 cell line, established from a rhabdomyosarcoma tumor (F Brasseur, unpublished data), was further confirmed by the absence of telomerase activity assessed by the telomeric repeat amplification protocol (Supplementary Figure 1). For the evaluation of subtelomeric DNA methylation levels, we chose to analyse the methylation status of the non-satellite D4Z4 subtelomeric repeats located at 4q35.2 and 10q26.3, as (1) the nucleotide sequence of the corresponding 3.3-kb D4Z4 repeats has been determined (Cacurri et al, 1998), (2) this region is known to be subjected to methylation on cytosine residues in normal tissues (van Overveld et al, 2003) and (3) the last D4Z4 repeat is mapped 23-24 kb from the 4q or 10q telomere in the current reference genome build (36.3) and further analyses indicated that D4Z4 is the last repetitive element in 4q and 10q (Riethman, 2008). Primers were designed to specifically amplify a 380-bp region of the D4Z4 repeat unit derived from 4q35.2 and 10q26.3 and containing 31 CpG dinucleotides.…”

Quality of life and pain in patients with facioscapulohumeral muscular dystrophy

“…In all patients, the clinical diagnosis had been confirmed by molecular testing, as described previously. 9,10 Pathological cut-off of the p13E11 EcoRI fragments was considered to be Յ40 kb in size. For each patient, the overall clinical severity was assessed using the Clinical Severity Scale, 30 which assigns scores ranging from 0.5 to 2 when the disease involves only facial and scapular muscles, and scores of 3 (mildly affected), 3.5 (moderately affected), and 4 -5 (severely affected, where a score of 5 is assigned to wheelchair-bound patients) when pelvic and lower limb muscles are involved.…”