Whole genome paired-end sequencing elucidates functional and phenotypic consequences of balanced chromosomal rearrangement in patients with developmental disorders

Abstract

BackgroundBalanced chromosomal rearrangements associated with abnormal phenotype are rare events, but may be challenging for genetic counselling, since molecular characterisation of breakpoints is not performed routinely. We used next-generation sequencing to characterise breakpoints of balanced chromosomal rearrangements at the molecular level in patients with intellectual disability and/or congenital anomalies.MethodsBreakpoints were characterised by a paired-end low depth whole genome sequencing (WGS) strategy and validated by Sanger sequencing. Expression study of disrupted and neighbouring genes was performed by RT-qPCR from blood or lymphoblastoid cell line RNA.ResultsAmong the 55 patients included (41 reciprocal translocations, 4 inversions, 2 insertions and 8 complex chromosomal rearrangements), we were able to detect 89% of chromosomal rearrangements (49/55). Molecular signatures at the breakpoints suggested that DNA breaks arose randomly and that there was no major influence of repeated elements. Non-homologous end-joining appeared as the main mechanism of repair (55% of rearrangements). A diagnosis could be established in 22/49 patients (44.8%), 15 by gene disruption (KANSL1, FOXP1, SPRED1, TLK2, MBD5, DMD, AUTS2, MEIS2, MEF2C, NRXN1, NFIX, SYNGAP1, GHR, ZMIZ1) and 7 by position effect (DLX5, MEF2C, BCL11B, SATB2, ZMIZ1). In addition, 16 new candidate genes were identified. Systematic gene expression studies further supported these results. We also showed the contribution of topologically associated domain maps to WGS data interpretation.ConclusionPaired-end WGS is a valid strategy and may be used for structural variation characterisation in a clinical setting.

Established and Novel Mechanisms Leading to de novo Genomic Rearrangements in the Human Germline

“…Such erroneous repair occurs at any position in the genome and often creates nonrecurrent deletions [Gu et al, 2008]. NHEJ-induced deletions were identified in patients with various congenital disorders [Schluth-Bolard et al, 2019]. Also, NHEJ leads to inversions, translocations, and duplications.…”

“…Also, NHEJ leads to inversions, translocations, and duplications. Indeed, most copy-number neutral nonrecurrent rearrangements, such as inversions and translocations, are predicted to be caused by NHEJ [Chiang et al, 2012;Schluth-Bolard et al, 2019].…”

Disruption of <b><i>PCDH10</i></b> and <b><i>TNRC18</i></b> Genes due to a Balanced Translocation

“…Different pathogenic mechanisms have been attributed as causes of the phenotypic alterations in BCAs, such as gene disruptions of dosage-sensitive genes at the breakpoints [Menten et al, 2006;Schneider et al, 2015], gene disruption followed by formation of chimeric genes expressing hybrid transcripts [Di Gregorio et al, 2013;Córdova-Fletes et al, 2015], and disruption of regulatory regions resulting in separation of cis-regulatory elements from the genes they control [Zepeda-Mendoza et al, 2018]. Precise breakpoint mapping is crucial for the genotypephenotype correlation [Redin et al, 2017;Schluth-Bolard et al, 2019] and can be an important approach to annotation of disease-associated genes. In addition to revealing pathogenic causes, the high-resolution breakpoint determination can give insights about the potential molecular mechanisms of chromosome break and repair and provide information into "mutational signatures" [Nilsson et al, 2017].…”

“…Additionally, chromosomal microarray-based genome-wide surveys are unable to further characterize balanced rearrangements. The evolution of accurate and affordable next-generation sequencing (NGS) techniques, such as whole genome sequencing (WGS), has been fundamental to elucidate breakpoints at nucleotide resolution contrib-uting to a molecular diagnosis [Aristidou et al, 2017[Aristidou et al, , 2018Schluth-Bolard et al, 2019]. Low coverage WGS, for example, is an efficient method for the detection and characterization of balanced translocations [Liang et al, 2017], and this characterization is improved when WGS is combined with cytogenomic methods [Moysés-Oliveira et al, 2019].…”

Whole-genome mate-pair sequencing of apparently balanced chromosome rearrangements reveals complex structural variations: two case studies

“…As a result, submicroscopic structural rearrangements could remain undetected, even when high-resolution banding technology is used. Molecular testing techniques such as FISH, CMA, and WGS have greatly improved the ability to identify ABCRs [6][7][8][9][10]. Recent studies have reported that WGS can detect extremely complex balance rearrangements, including genome structural variations at the molecular level, which otherwise remain detected by conventional G banding techniques [11].…”