Abstract
Foldback inversions, also called inverted duplications, have been observed in human genetic diseases and cancers. Here, we used a Saccharomyces cerevisiae genetic system that generates gross chromosomal rearrangements (GCRs) mediated by foldback inversions combined with whole-genome sequencing to study their formation. Foldback inversions were mediated by formation of single-stranded DNA hairpins. Two types of hairpins were identified: small-loop hairpins that were suppressed by MRE11, SAE2, SLX1, and YKU80 and large-loop hairpins that were suppressed by YEN1, TEL1, SWR1, and MRC1. Analysis of CRISPR/Cas9-induced double strand breaks (DSBs) revealed that long-stem hairpin-forming sequences could form foldback inversions when proximal or distal to the DSB, whereas short-stem hairpin-forming sequences formed foldback inversions when proximal to the DSB. Finally, we found that foldback inversion GCRs were stabilized by secondary rearrangements, mostly mediated by different homologous recombination mechanisms including single-strand annealing; however, POL32-dependent break-induced replication did not appear to be involved forming secondary rearrangements.
Highlights
Organisms invest substantial effort into properly replicating their genomes and preventing DNA damage from leading to the accumulation of mutations and gross chromosomal rearrangements (GCRs), such as translocations, deletions and inversions
Thick-hashed arrows underneath the plot indicate the connectivity between the portions of the GCR that map to the different regions of the reference chromosome. (E) Example read depth plot for chrV and chrXIV from a foldback inversion GCR resolved by a ura3-52/YNLCTy1-1 homology-mediated rearrangement displayed as in panel D, showing the duplication of the region of chrXIV between TEL14L and YNLCTy1-1
These inversions arose by a mechanism whereby a terminal region of chrV L was broken, a region on the centromeric side of the break was duplicated in inverted orientation by a mechanism involving a hairpin junction with a large intervening loop, and the other end of the duplicated region was joined to the telomeric terminal fragment of chrV L, capturing a telomere
Summary
Organisms invest substantial effort into properly replicating their genomes and preventing DNA damage from leading to the accumulation of mutations and gross chromosomal rearrangements (GCRs), such as translocations, deletions and inversions. Considerable recent progress has been made through the discovery of cancer susceptibility syndromes associated with increased rates of accumulation of mutations and GCRs as well as the use of whole genome sequencing (WGS) to characterize cancer genomes and to identify genome instability signatures such as chromothripsis and chromoplexy (Baca et al, 2013; Li et al, 2020; Meyerson and Pellman, 2011). In spite of this progress, a comprehensive understanding of the pathways and mechanisms that suppress or promote the formation of GCRs in mammalian cells is not yet available
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