In this study, we show that evolutionarily conserved chromosome loop anchors bound by CCCTC-binding factor (CTCF) and cohesin are vulnerable to DNA double strand breaks (DSBs) mediated by ...topoisomerase 2B (TOP2B). Polymorphisms in the genome that redistribute CTCF/cohesin occupancy rewire DNA cleavage sites to novel loop anchors. While transcription- and replication-coupled genomic rearrangements have been well documented, we demonstrate that DSBs formed at loop anchors are largely transcription-, replication-, and cell-type-independent. DSBs are continuously formed throughout interphase, are enriched on both sides of strong topological domain borders, and frequently occur at breakpoint clusters commonly translocated in cancer. Thus, loop anchors serve as fragile sites that generate DSBs and chromosomal rearrangements.
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•Chromosome loop anchors bound by CTCF and Cohesin are vulnerable to DNA breaks•Breaks are transcription independent, mediated by TOP2B and correlate with cohesin•Translocation breakpoint regions in various cancers are enriched at loop anchors•Chromosome folding and topological stress relief go hand in hand
Chromatin assembly into higher-order structures generates torsional stress that makes chromosome loop anchor regions more vulnerable to topoisomerase 2-mediated DNA breaks.
Cells deficient in the Brca1 and Brca2 genes have reduced capacity to repair DNA double-strand breaks by homologous recombination and consequently are hypersensitive to DNA-damaging agents, including ...cisplatin and poly(ADP-ribose) polymerase (PARP) inhibitors. Here we show that loss of the MLL3/4 complex protein, PTIP, protects Brca1/2-deficient cells from DNA damage and rescues the lethality of Brca2-deficient embryonic stem cells. However, PTIP deficiency does not restore homologous recombination activity at double-strand breaks. Instead, its absence inhibits the recruitment of the MRE11 nuclease to stalled replication forks, which in turn protects nascent DNA strands from extensive degradation. More generally, acquisition of PARP inhibitors and cisplatin resistance is associated with replication fork protection in Brca2-deficient tumour cells that do not develop Brca2 reversion mutations. Disruption of multiple proteins, including PARP1 and CHD4, leads to the same end point of replication fork protection, highlighting the complexities by which tumour cells evade chemotherapeutic interventions and acquire drug resistance.
Nonreciprocal translocations and gene amplifications are commonly found in human tumors. Although little is known about the mechanisms leading to such aberrations, tissue culture models predict that ...they can arise from DNA breakage, followed by cycles of chromatid fusion, asymmetric mitotic breakage, and replication. Mice deficient in both a nonhomologous end joining (NHEJ) DNA repair protein and the p53 tumor suppressor develop lymphomas at an early age harboring amplification of an IgH/c-myc fusion. Here we report that these chromosomal rearrangements are initiated by a recombination activating gene (RAG)-induced DNA cleavage. Subsequent DNA repair events juxtaposing IgH and c-myc are mediated by a break-induced replication pathway. Cycles of breakage-fusion-bridge result in amplification of IgH/c-myc while chromosome stabilization occurs through telomere capture. Thus, mice deficient in NHEJ provide excellent models to study the etiology of unbalanced translocations and amplification events during tumorigenesis.
Poly-(ADP-ribose) polymerase (PARP) inhibitors (PARPis) selectively kill BRCA1/2-deficient cells, but their efficacy in BRCA-deficient patients is limited by drug resistance. Here, we used derived ...cell lines and cells from patients to investigate how to overcome PARPi resistance. We found that the functions of BRCA1 in homologous recombination (HR) and replication fork protection are sequentially bypassed during the acquisition of PARPi resistance. Despite the lack of BRCA1, PARPi-resistant cells regain RAD51 loading to DNA double-stranded breaks (DSBs) and stalled replication forks, enabling two distinct mechanisms of PARPi resistance. Compared with BRCA1-proficient cells, PARPi-resistant BRCA1-deficient cells are increasingly dependent on ATR for survival. ATR inhibitors (ATRis) disrupt BRCA1-independent RAD51 loading to DSBs and stalled forks in PARPi-resistant BRCA1-deficient cells, overcoming both resistance mechanisms. In tumor cells derived from patients, ATRis also overcome the bypass of BRCA1/2 in fork protection. Thus, ATR inhibition is a unique strategy to overcome the PARPi resistance of BRCA-deficient cancers.
Histone H2AX becomes phosphorylated in chromatin domains flanking sites of DNA double-strand breakage associated with γ-irradiation, meiotic recombination, DNA replication, and antigen receptor ...rearrangements. Here, we show that loss of a single
H2AX allele compromises genomic integrity and enhances the susceptibility to cancer in the absence of p53. In comparison with heterozygotes, tumors arise earlier in the H2AX homozygous null background, and
H2AX
−/−
p53
−/− lymphomas harbor an increased frequency of clonal nonreciprocal translocations and amplifications. These include complex rearrangements that juxtapose the c-
myc oncogene to antigen receptor loci. Restoration of the
H2AX null allele with wild-type
H2AX restores genomic stability and radiation resistance, but this effect is abolished by substitution of the conserved serine phosphorylation sites in H2AX with alanine or glutamic acid residues. Our results establish
H2AX as genomic caretaker that requires the function of both gene alleles for optimal protection against tumorigenesis.
Burning bridges in cancer genomes Paiano, Jacob; Nussenzweig, André
Science (American Association for the Advancement of Science),
04/2020, Letnik:
368, Številka:
6488
Journal Article
Recenzirano
Cytoskeletal forces break chromosomal fusions and trigger mutational avalanches
Cancer is driven by mutations. A large fraction of mutations arise from unavoidable errors of DNA replication, which ...accumulate gradually over generations (
1
). Recent studies indicate that cancer genomes frequently harbor mutation signatures that result from chromothripsis. This is a sudden form of localized and massive chromosomal rearrangement, usually involving one or a few chromosomes. Chromothripsis is thought to arise through a single shattering event, after which tens to hundreds of chromosomal segments are joined in a random order and orientation. This one-off catastrophic event is common in cancer, with frequencies reaching 65% in certain cancer types (
2
). Until now, the mechanisms that explain this burst of mutagenesis remained largely unclear. On page 282 of this issue, Umbreit
et al.
(
3
) provide a unifying account of how extreme genome complexity is an outcome of a mutagenesis mechanism that involves aberrant DNA replication in human cells.
CTCF is a barrier for 2C-like reprogramming Olbrich, Teresa; Vega-Sendino, Maria; Tillo, Desiree ...
Nature communications,
08/2021, Letnik:
12, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Totipotent cells have the ability to generate embryonic and extra-embryonic tissues. Interestingly, a rare population of cells with totipotent-like potential, known as 2 cell (2C)-like cells, has ...been identified within ESC cultures. They arise from ESC and display similar features to those found in the 2C embryo. However, the molecular determinants of 2C-like conversion have not been completely elucidated. Here, we show that the CCCTC-binding factor (CTCF) is a barrier for 2C-like reprogramming. Indeed, forced conversion to a 2C-like state by the transcription factor DUX is associated with DNA damage at a subset of CTCF binding sites. Depletion of CTCF in ESC efficiently promotes spontaneous and asynchronous conversion to a 2C-like state and is reversible upon restoration of CTCF levels. This phenotypic reprogramming is specific to pluripotent cells as neural progenitor cells do not show 2C-like conversion upon CTCF-depletion. Furthermore, we show that transcriptional activation of the ZSCAN4 cluster is necessary for successful 2C-like reprogramming. In summary, we reveal an unexpected relationship between CTCF and 2C-like reprogramming.
53BP1 is a DNA damage protein that forms phosphorylated H2AX (γ-H2AX) dependent foci in a 1 Mb region surrounding DNA double-strand breaks (DSBs). In addition, 53BP1 promotes genomic stability by ...regulating the metabolism of DNA ends. We have compared the joining rates of paired DSBs separated by 1.2 kb to 27 Mb on chromosome 12 in the presence or absence of 53BP1. 53BP1 facilitates joining of intrachromosomal DSBs but only at distances corresponding to γ-H2AX spreading. In contrast, DNA end protection by 53BP1 is distance independent. Furthermore, analysis of 53BP1 mutants shows that chromatin association, oligomerization, and N-terminal ATM phosphorylation are all required for DNA end protection and joining as measured by immunoglobulin class switch recombination. These data elucidate the molecular events that are required for 53BP1 to maintain genomic stability and point to a model wherein 53BP1 and H2AX cooperate to repress resection of DSBs.
► 53BP1 facilitates the joining of DSBs depending on the distance between breaks ► 53BP1 prevents DNA end resection independent of the distance between breaks ► In the absence of H2AX, 53BP1 is chromatin associated but does not block resection ► Multiple functions of 53BP1 are needed for end protection and Ig class switching
Chromosomal translocations between antigen receptor loci and oncogenes are a hallmark of lymphoid cancers. Several new studies now reveal that programmed DNA breaks created during assembly of antigen ...receptor genes can be channeled into an alternative DNA end-joining pathway that is implicated in the chromosomal translocations of lymphoid cancers (
Corneo et al., 2007; Soulas-Sprauel et al., 2007; Yan et al., 2007).
Replication origins, fragile sites, and rDNA have been implicated as sources of chromosomal instability. However, the defining genomic features of replication origins and fragile sites are among the ...least understood elements of eukaryote genomes. Here, we map sites of replication initiation and breakage in primary cells at high resolution. We find that replication initiates between transcribed genes within nucleosome-depleted structures established by long asymmetrical poly(dA:dT) tracts flanking the initiation site. Paradoxically, long (>20 bp) (dA:dT) tracts are also preferential sites of polar replication fork stalling and collapse within early-replicating fragile sites (ERFSs) and late-replicating common fragile sites (CFSs) and at the rDNA replication fork barrier. Poly(dA:dT) sequences are fragile because long single-strand poly(dA) stretches at the replication fork are unprotected by the replication protein A (RPA). We propose that the evolutionary expansion of poly(dA:dT) tracts in eukaryotic genomes promotes replication initiation, but at the cost of chromosome fragility.
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•Genome-wide map of DNA breaks due to replication stress in mammalian cells•Poly(dA:dT) tracts are natural polar replication barriers and fragile sites•Common mechanism for DNA breakage at early- and late-replicating fragile sites•AT richness and poly(dA:dT) motifs are determinants of origin usage in mammals
Mammalian replication origins are fragile sites defined by poly-dA/dT stretches that are nucleosome free and devoid of the single-strand DNA-protecting protein RPA.
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