Non Random Distribution of DMD Deletion Breakpoints and Implication of Double Strand Breaks Repair and Replication Error Repair Mechanisms

Marey, Isabelle; Yaou, Rabah Ben; Deburgrave, Nathalie; Vasson, Aurélie; Nectoux, Juliette; Leturcq, F.; Eymard, B.; Laforêt, Pascal; Béhin, Anthony; Stojkovic, Tanya; Mayer, M.; Tiffreau, V.; Desguerre, Isabelle; Boyer, Françoise; Nadaj‐Pakleza, Aleksandra; Ferrer, Xavier; Wahbi, Karim; Bécane, Henri-Marc; Claustres, Mireille; Chelly, Jamel; Cossée, Mireille

doi:10.3233/jnd-150134

Cited by 7 publications

(11 citation statements)

References 48 publications

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“…Intron 44 spans the largest length (12%) of all introns in the DMD gene, and harbors more than 25% of the deletion start points ( Tong et al, 2020 ). However, only a few previous small-sample studies have focused on the fine structure of intron 44 ( Blonden et al, 1991 ; Love et al, 1991 ; Miyazaki et al, 2009 ; Marey et al, 2016 ). The main difficulty for the sequence analysis of intron 44 may be that short-read sequencing is not adept at locating the exact breakpoints in such a large intron with tremendous repetitive regions.…”

Section: Discussionmentioning

confidence: 99%

“…Not to mention that many studies only performed MLPA and did not search for breakpoints. Recently, Marey et al (2016) mapped 39 patients with deletions starting from intron 44 based on an aCGH array, and found that 48.7% of breakpoints clustered in the distal 50 kb regions and 33.3% were clustered in regions less than 700 bp. Researchers also found that repetitive elements, and palindromic and T-A sequences were present in the vicinity of the breakpoints ( Toffolatti et al, 2002 ; Marey et al, 2016 ).…”

Section: Introductionmentioning

confidence: 99%

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Sequence and Structure Characteristics of 22 Deletion Breakpoints in Intron 44 of the DMD Gene Based on Long-Read Sequencing

et al. 2021

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Purpose: Exon deletions make up to 80% of mutations in the DMD gene, which cause Duchenne and Becker muscular dystrophy. Exon 45-55 regions were reported as deletion hotspots and intron 44 harbored more than 25% of deletion start points. We aimed to investigate the fine structures of breakpoints in intron 44 to find potential mechanisms of large deletions in intron 44.Methods: Twenty-two dystrophinopathy patients whose deletion started in intron 44 were sequenced using long-read sequencing of a DMD gene capture panel. Sequence homology, palindromic sequences, and polypyrimidine sequences were searched at the breakpoint junctions. RepeatMasker was used to analyze repetitive elements and Mfold was applied to predict secondary DNA structure.Results: With a designed DMD capture panel, 22 samples achieved 2.25 gigabases and 1.28 million reads on average. Average depth was 308× and 99.98% bases were covered at least 1×. The deletion breakpoints in intron 44 were scattered and no breakpoints clustered in any region less than 500 bp. A total of 72.7% of breakpoints located in distal 100 kb of intron 44 and more repetitive elements were found in this region. Microhomologies of 0–1 bp were found in 36.4% (8/22) of patients, which corresponded with non-homologous end-joining. Microhomologies of 2–20 bp were found in 59.1% (13/22) of patients, which corresponded with microhomology-mediated end-joining. Moreover, a 7 bp insertion was found in one patient, which might be evidence of aberrant replication origin firing. Palindromic sequences, polypyrimidine sequences, and small hairpin loops were found near several breakpoint junctions. No evidence of large hairpin loop formation in deletion root sequences was observed.Conclusion: This study was the first to explore possible mechanisms underlying exon deletions starting from intron 44 of the DMD gene based on long-read sequencing. Diverse mechanisms might be associated with deletions in the DMD gene.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Sequence and Structure Characteristics of 22 Deletion Breakpoints in Intron 44 of the DMD Gene Based on Long-Read Sequencing

et al. 2021

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“…The selected flanking reads analysis in intron 44 found that microhomology or extended microhomology sequences existed around the junction reads, and one subject (D37) without microhomology sequences had long motifs (Tables and ). The above results illustrate that microhomology (Marey et al, ) and long motifs might contribute to the instability of intron 44. We further performed flanking sequence analysis around the breakpoints in all of the 37 patients, and the results illustrated that DMD gene instability did not arise from a single underlying cause.…”

Section: Discussionmentioning

confidence: 55%

“…MMRDR was hypothesized to cause replication fork stalling and template switching, which could induce complex deletion and duplication rearrangement (Lee, Carvalho, & Lupski, 2007 (Roth, Porter, & Wilson, 1985). In the present study, some of the 200 flanking regions of the breakpoints were observed to be palindromic (CTTC) and contained low-copy repeat sequences (AAAA), which could promote DNA instability (Ankala et al, 2012;Marey et al, 2016;Sen et al, 2006) and then produce a direct ligation. Additionally, small-sequence insertions and deletions were frequently found (in 14% of patients with DMD/BMD), which could be explained by repair synthesis primed by terminal homologies.…”

Section: Discussionmentioning

confidence: 61%

Exonic rearrangements in DMD in Chinese Han individuals affected with Duchenne and Becker muscular dystrophies

Ling

Dai

et al. 2019

Human Mutation

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Exonic deletions and duplications within DMD are the main pathogenic variants in Duchenne and Becker muscular dystrophies (DMD/BMD). However, few studies have profiled the flanking sequences of breakpoints and the potential mechanism underlying the breakpoints in different fragile regions of DMD. In this study, 896 Chinese male probands afflicted with DMD/BMD were selected from unrelated families and analyzed using multiplex ligation-dependent probe amplification of the DMD gene, in which we identified exon deletions in 784 subjects and duplications in 112 subjects. Deletions occurred most frequently in the genomic region encompassing exons 45-55,accounting for 73% of all deletion patterns. Furthermore, to unravel the potential mechanism that induced breaks, DMD gene capture and sequencing were performed to identify the breakpoints in 37 subjects with deletions encompassing exons 45-55 of DMD; we found that DMD instability did not arise from a single cause; instead, longsequence motifs, nonconsensus microhomologies, low-copy repeats, and microindels were embedded around the breakpoints, which may predispose DMD to instability. In summary, this study highlights the heterogeneous characteristics of the flanking sequences around the breakpoints and helps us to understand the mechanism underlying DMD gene instability.

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“…Indeed the 1, 7, and 44 giant introns are just downstream of out-of-frame exons (1, 7, and 44), which are (not surprisingly) the DMD deletion hot spots. These three introns are also very rich in short tandem repeats (especially TTTAAA or (T)n(A) n ), which are known to cause aberrant firing of the replication fork, as frequent mechanisms responsible for micro-homology related recombination events (Marey et al, 2016). We would like to underline that these three exons are the upstream exons of regions encoding extremely important DYS domains: the actin binding domain (exon 1, although possibly dispensable, as suggested by Wein et al, 2014), the Hinge 1 (exon 7) and the nNOS binding domain, involved in DYS signaling functions (Rybakova et al, 2000).…”

Section: Dmd Deletions and Duplications: Frequency Distribution Topmentioning

confidence: 99%

The Genetic Landscape of Dystrophin Mutations in Italy: A Nationwide Study

Neri¹,

Rossi²,

Trabanelli³

et al. 2020

Front. Genet.

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in 57%, duplications in 11% and small mutations in 32%. In BMD, we found deletions in 78%, duplications in 9% and small mutations in 13%. In BMD, there are a higher number of deletions, and small mutations are more frequent than duplications. Among small mutations that are generally frequent in both phenotypes, 44% of DMD and 36% of BMD are nonsense, thus, eligible for stop codon read-through therapy; 63% of all out-of-frame deletions are eligible for single exon skipping. Patients were also assigned to Italian regions and showed interesting regional differences in mutation distribution. The full genetic characterization in this large, nationwide cohort has allowed us to draw several correlations between DMD/BMD genotype landscapes and mutation frequency, mutation types, mutation locations along the gene, exon/intron architecture, and relevant protein d o m a i n , w i t h e ff e c t s on p o p u l a t i o n ge n e t i c c h a r a c t e r i s t i c s a n d ne w personalized therapies.

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Non Random Distribution of DMD Deletion Breakpoints and Implication of Double Strand Breaks Repair and Replication Error Repair Mechanisms

Cited by 7 publications

References 48 publications

Sequence and Structure Characteristics of 22 Deletion Breakpoints in Intron 44 of the DMD Gene Based on Long-Read Sequencing

Sequence and Structure Characteristics of 22 Deletion Breakpoints in Intron 44 of the DMD Gene Based on Long-Read Sequencing

Exonic rearrangements in DMD in Chinese Han individuals affected with Duchenne and Becker muscular dystrophies

The Genetic Landscape of Dystrophin Mutations in Italy: A Nationwide Study

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