Identification and analysis of deletion breakpoints in four Mohr-Tranebjærg syndrome (MTS) patients

Mohr-Tranebjærg syndrome is an X-linked syndrome characterized by sensorineural hearing impairment in childhood, followed by progressive neurodegeneration leading to a broad phenotypic spectrum. Genetically MTS is caused by pathogenic variants in the TIMM8A gene, including gene deletions and larger contiguous gene deletions. Some of the latter involve the neighboring gene BTK, resulting in agammaglobulinemia. By next-generation mate-pair sequencing we have mapped the chromosomal deletion breakpoints of one MTS case and three XLA-MTS cases and used breakpoint-spanning PCR to fine map the breakpoints by Sanger sequencing. Two of the XLA-MTS cases presented with large deletions (63.5 and 27.2 kb), and the junctional regions were characterized by long stretches of microhomology, indicating that the events have emerged through homologous recombination. Conversely, the MTS case exhibited a small 2 bp region of microhomology, and the regions were not characterized by extensive microhomology. The third XLA-MTS case had a more complex breakpoint, including a 59 bp inverted insertion, thus at least four breakpoints were involved in this event. In conclusion, mate-pair library generation combined with next-generation sequencing is an efficient method for breakpoint identification, also in regions characterized by repetitive elements

Balanced chromosomal rearrangements offer insights into coding and noncoding genomic features associated with developmental disorders

Balanced chromosomal rearrangements (BCRs), including inversions, translocations, and insertions, reorganize large sections of the genome and contribute substantial risk for developmental disorders (DDs). However, the rarity and lack of systematic screening for BCRs in the population has precluded unbiased analyses of the genomic features and mechanisms associated with risk for DDs versus normal developmental outcomes. Here, we sequenced and analyzed 1,420 BCR breakpoints across 710 individuals, including 406 DD cases and the first large-scale collection of 304 control BCR carriers. We found that BCRs were not more likely to disrupt genes in DD cases than controls, but were seven-fold more likely to disrupt genes associated with dominant DDs (21.3% of cases vs. 3.4% of controls; P = 1.60×10$^{−12}$). Moreover, BCRs that did not disrupt a known DD gene were significantly enriched for breakpoints that altered topologically associated domains (TADs) containing dominant DD genes in cases compared to controls (odds ratio [OR] = 1.43, P = 0.036). We discovered six TADs enriched for noncoding BCRs (false discovery rate < 0.1) that contained known DD genes (MEF2C, FOXG1, SOX9, BCL11A, BCL11B, and SATB2) and represent candidate pathogenic long-range positional effect (LRPE) loci. These six TADs were collectively disrupted in 7.4% of the DD cohort. Phased Hi-C analyses of five cases with noncoding BCR breakpoints localized to one of these putative LRPEs, the 5q14.3 TAD encompassing MEF2C, confirmed extensive disruption to local 3D chromatin structures and reduced frequency of contact between the MEF2C promoter and annotated enhancers. We further identified six genomic features enriched in TADs preferentially disrupted by noncoding BCRs in DD cases versus controls and used these features to build a model to predict TADs at risk for LRPEs across the genome. These results emphasize the potential impact of noncoding structural variants to cause LRPEs in unsolved DD cases, as well as the complex interaction of features associated with predicting three-dimensional chromatin structures intolerant to disruption

A germline chromothripsis event stably segregating in 11 individuals through three generations

Purpose: Parentally transmitted germ-line chromothripsis (G-CTH) has been identified in only a few cases. Most of these rearrangements were stably transmitted, in an unbalanced form, from a healthy mother to her child with congenital abnormalities probably caused by de novo copy-number changes of dosage sensitive genes. We describe a G-CTH transmitted through three generations in 11 healthy carriers. Methods: Conventional cytogenetic analysis, mate-pair sequencing, and polymerase chain reaction (PCR) were used to identify the chromosome rearrangement and characterize the breakpoints in all three generations. Results: We identified an apparently balanced translocation t(3;5), later shown to be a G-CTH, in all individuals of a three-generation family. The G-CTH stably segregated without occurrence of additional rearrangements; however, several spontaneous abortions were reported, possibly due to unbalanced transmission. Although seven protein-coding genes are interrupted, no clinical features can be definitively attributed to the affected genes. However, it can be speculated that truncation of one of these genes, encoding ataxia–telangiectasia and Rad3-related protein kinase (ATR), a key component of the DNA damage response, may be related to G-CTH formation. Conclusion: G-CTH rearrangements are not always associated with abnormal phenotypes and may be misinterpreted as balanced two-way translocations, suggesting that G-CTH is an underdiagnosed phenomenon

Identification and analysis of deletion breakpoints in four Mohr-Tranebjærg syndrome (MTS) patients

Mohr-Tranebjærg syndrome is an X-linked syndrome characterized by sensorineural hearing impairment in childhood, followed by progressive neurodegeneration leading to a broad phenotypic spectrum. Genetically MTS is caused by pathogenic variants in the TIMM8A gene, including gene deletions and larger contiguous gene deletions. Some of the latter involve the neighboring gene BTK, resulting in agammaglobulinemia. By next‐generation mate‐pair sequencing we have mapped the chromosomal deletion breakpoints of one MTS case and three XLA-MTS cases and used breakpoint-spanning PCR to fine map the breakpoints by Sanger sequencing. Two of the XLA-MTS cases presented with large deletions (63.5 and 27.2&nbsp;kb), and the junctional regions were characterized by long stretches of microhomology, indicating that the events have emerged through homologous recombination. Conversely, the MTS case exhibited a small 2&nbsp;bp region of microhomology, and the regions were not characterized by extensive microhomology. The third XLA-MTS case had a more complex breakpoint, including a 59&nbsp;bp inverted insertion, thus at least four breakpoints were involved in this event. In conclusion, mate-pair library generation combined with next-generation sequencing is an efficient method for breakpoint identification, also in regions characterized by repetitive elements

A germline chromothripsis event stably segregating in 11 individuals through three generations

Purpose: Parentally transmitted germ-line chromothripsis (G-CTH) has been identified in only a few cases. Most of these rearrangements were stably transmitted, in an unbalanced form, from a healthy mother to her child with congenital abnormalities probably caused by de novo copy-number changes of dosage sensitive genes. We describe a G-CTH transmitted through three generations in 11 healthy carriers. Methods: Conventional cytogenetic analysis, mate-pair sequencing, and polymerase chain reaction (PCR) were used to identify the chromosome rearrangement and characterize the breakpoints in all three generations. Results: We identified an apparently balanced translocation t(3;5), later shown to be a G-CTH, in all individuals of a three-generation family. The G-CTH stably segregated without occurrence of additional rearrangements; however, several spontaneous abortions were reported, possibly due to unbalanced transmission. Although seven protein-coding genes are interrupted, no clinical features can be definitively attributed to the affected genes. However, it can be speculated that truncation of one of these genes, encoding ataxia-telangiectasia and Rad3-related protein kinase (ATR), a key component of the DNA damage response, may be related to G-CTH formation. Conclusion: G-CTH rearrangements are not always associated with abnormal phenotypes and may be misinterpreted as balanced two-way translocations, suggesting that G-CTH is an underdiagnosed phenomenon