Large Inverted Duplications in the Human Genome Form via a Fold-Back Mechanism

Abstract

Inverted duplications are a common type of copy number variation (CNV) in germline and somatic genomes. Large duplications that include many genes can lead to both neurodevelopmental phenotypes in children and gene amplifications in tumors. There are several models for inverted duplication formation, most of which include a dicentric chromosome intermediate followed by breakage-fusion-bridge (BFB) cycles, but the mechanisms that give rise to the inverted dicentric chromosome in most inverted duplications remain unknown. Here we have combined high-resolution array CGH, custom sequence capture, next-generation sequencing, and long-range PCR to analyze the breakpoints of 50 nonrecurrent inverted duplications in patients with intellectual disability, autism, and congenital anomalies. For half of the rearrangements in our study, we sequenced at least one breakpoint junction. Sequence analysis of breakpoint junctions reveals a normal-copy disomic spacer between inverted and non-inverted copies of the duplication. Further, short inverted sequences are present at the boundary of the disomic spacer and the inverted duplication. These data support a mechanism of inverted duplication formation whereby a chromosome with a double-strand break intrastrand pairs with itself to form a “fold-back” intermediate that, after DNA replication, produces a dicentric inverted chromosome with a disomic spacer corresponding to the site of the fold-back loop. This process can lead to inverted duplications adjacent to terminal deletions, inverted duplications juxtaposed to translocations, and inverted duplication ring chromosomes.

Organelle DNA rearrangement mapping reveals U-turn-like inversions as a major source of genomic instability in Arabidopsis and humans

“…In this regard, it will be interesting to evaluate if particular patterns of mitochondrial genomic instability are observed in the context of these disorders. Genomic rearrangements associated with inversions in the human nuclear genome have also been linked to leukemia, autism, and intellectual disability (Pui et al 1992;Hermetz et al 2014). It would therefore also be interesting to determine whether such rearrangements also occur through a U-turn-like mechanism and lead to the onset of such phenotypes.…”

Next-Generation Sequencing of Duplication CNVs Reveals that Most Are Tandem and Some Create Fusion Genes at Breakpoints

“…This type of rearrangement forms via a distinct mechanism involving a dicentric chromosome intermediate that breaks to give rise to a characteristic terminal deletion adjacent to inverted duplications separated by a short disomic spacer. 33 Another duplication that appeared to be terminal by microarray analysis turned out to be in direct orientation. WGS of EGL825's terminal duplication suggested an interchromosomal duplication between chromosomes 9 and 12 (Table S2); however, FISH confirmed that this is an intrachromosomal duplication of the short arm of chromosome 9.…”

“…Inverted duplications adjacent to terminal deletions have a characteristic appearance via microarray analysis and in almost all cases occur de novo. 33,63 Terminal gains are most often unbalanced translocations, but in rare cases might be inverted or direct intrachromosomal duplications. Chromosomes with a terminal duplication of one end and a terminal deletion of the other end can be generated by recombination within an inversion loop.…”

Inverted duplication with deletion: First interstitial case suggesting a novel undescribed mechanism of formation

“…The main mechanism for inverted duplication formation was originally believed to be a double strand break leading to telomere loss and fusion of sister chromatids (U-type exchange) followed by breakage of the dicentric chromosome and breakage-fusion-bridge cycles [Knijnenburg et al, 2007]. Recently, a fold-back mechanism has been described whereby after the initial double strand break and telomere loss single-stranded DNA folds back and intra-strand pairs resulting in an intrachromosomal inverted duplication [Hermetz et al, 2014]. This same mechanism has been put forward to explain the formation of inverted duplication on acentric markers [Murmann et al, 2009].…”

“…A clear distinction can be made between these two mechanisms through the identification of a single copy region between the deleted and duplicated regions in case of NAHR based rearrangement. Through the use of next generation sequencing, Hermetz et al [2014] proposed a new mechanism of formation based on an intrastrand fold-back of a broken DNA that anneals to itself at site of inverted sequence homology, which may represent the leading pathway to generation of the dicentric.…”