Sister chromatid telomere fusions, but not NHEJ-mediated inter-chromosomal telomere fusions, occur independently of DNA ligases 3 and 4

Abstract

Telomeres shorten with each cell division and can ultimately become substrates for nonhomologous end-joining repair, leading to large-scale genomic rearrangements of the kind frequently observed in human cancers. We have characterized more than 1400 telomere fusion events at the single-molecule level, using a combination of high-throughput sequence analysis together with experimentally induced telomeric double-stranded DNA breaks. We show that a single chromosomal dysfunctional telomere can fuse with diverse nontelomeric genomic loci, even in the presence of an otherwise stable genome, and that fusion predominates in coding regions. Fusion frequency was markedly increased in the absence of TP53 checkpoint control and significantly modulated by the cellular capacity for classical, versus alternative, nonhomologous end joining (NHEJ). We observed a striking reduction in inter-chromosomal fusion events in cells lacking DNA ligase 4, in contrast to a remarkably consistent profile of intra-chromosomal fusion in the context of multiple genetic knockouts, including DNA ligase 3 and 4 doubleknockouts. We reveal distinct mutational signatures associated with classical NHEJ-mediated inter-chromosomal, as opposed to alternative NHEJ-mediated intra-chromosomal, telomere fusions and evidence for an unanticipated sufficiency of DNA ligase 1 for these intra-chromosomal events. Our findings have implications for mechanisms driving cancer genome evolution.

Extensive telomere erosion is consistent with localised clonal expansions in Barrett’s metaplasia

“…Thus if telomere dysfunction is driving copy number changes at specific chromosome arms then this is not related to the absolute length of the associated telomere. Instead this is more consistent with a situation in which telomere erosion across all chromosome ends leads to the accumulation of short dysfunctional telomeres that are capable of fusion and driving genome-wide copy number changes and selection for specific chromosomal rearrangements [38]. Whilst patient matched genomic data from Barrett’s metaplasia and oesophageal adenocarcinoma indicated a comparatively low level of copy number changes in Barrett’s metaplasia, there was a clear increase in copy number changes within adenocarcinomas [39]; these types of genomic mutation may be consistent with telomere dysfunction during the transition from Barrett’s metaplasia to oesophageal carcinoma.…”

Telomeres in cancer: tumour suppression and genome instability

“…Non-reciprocal trans-locations occur during tumorigenesis in mice with shortening telomeres 1 and are a frequent class of rearrangements in cancer 89 . Sequence analysis of more than 1,000 telomere fusion events has shown that a chromosome end lacking telomere protection can recombine with diverse chromosome-internal loci 90 .…”

A Single Defined Sister Chromatid Fusion Destabilizes Cell Cycle through Micronuclei Formation

“…The preference of 154 large truncation and microhomology at the fusion junction suggests that sister chromatid fusion 155 is processed by microhomology-mediated end joining (MMEJ), which is active in late S/G2 156 phase when sister chromatids are present [30]. This profile of the fusion junction is consistent 157 with naturally occurred and TALEN-induced chromosome end-to-end 158 fusions [31] [32] [33] [21]. 159 Construction of the control system 160 Since the XpSIS relies on targeting of the subtelomeric cassette by CRISPR/Cas9 and the 161 resulting expression of mCitrine, we designed a control system, in which CRISPR/Cas9 162 targeting of Xp subtelomere induces mCitrine expression without sister chromatid fusion ( Figure 163 2A, B).…”