The elusive evidence for chromothripsis

The chromothripsis hypothesis suggests an extraordinary one-step catastrophic genomic event allowing a chromosome to ‘shatter into many pieces’ and reassemble into a functioning chromosome. Recent efforts have aimed to detect chromothripsis by looking for a genomic signature, characterized by a large number of breakpoints (50–250), but a limited number of oscillating copy number states (2–3) confined to a few chromosomes. The chromothripsis phenomenon has become widely reported in different cancers, but using inconsistent and sometimes relaxed criteria for determining rearrangements occur simultaneously rather than progressively. We revisit the original simulation approach and show that the signature is not clearly exceptional, and can be explained using only progressive rearrangements. For example, 3.9% of progressively simulated chromosomes with 50–55 breakpoints were dominated by two or three copy number states. In addition, by adjusting the parameters of the simulation, the proposed footprint appears more frequently. Lastly, we provide an algorithm to find a sequence of progressive rearrangements that explains all observed breakpoints from a proposed chromothripsis chromosome. Thus, the proposed signature cannot be considered a sufficient proof for this extraordinary hypothesis. Great caution should be exercised when labeling complex rearrangements as chromothripsis from genome hybridization and sequencing experiments.     

Chromosome catastrophes involve replication mechanisms generating complex genomic rearrangements

Complex genomic rearrangements (CGRs) consisting of two or more breakpoint junctions have been observed in genomic disorders. Recently, a chromosome catastrophe phenomenon termed chromothripsis, in which numerous genomic rearrangements are apparently acquired in one single catastrophic event, was described in multiple cancers. Here, we show that constitutionally acquired CGRs share similarities with cancer chromothripsis. In the 17 CGR cases investigated, we observed localization and multiple copy number changes including deletions, duplications, and/or triplications, as well as extensive translocations and inversions. Genomic rearrangements involved varied in size and complexities; in one case, array comparative genomic hybridization revealed 18 copy number changes. Breakpoint sequencing identified characteristic features, including small templated insertions at breakpoints and microhomology at breakpoint junctions, which have been attributed to replicative processes. The resemblance between CGR and chromothripsis suggests similar mechanistic underpinnings. Such chromosome catastrophic events appear to reflect basic DNA metabolism operative throughout an organism's life cycle.     

Chromothripsis: Breakage-fusion-bridge over and over again

The acquisition of massive but localized chromosome translocations, a phenomenon termed chromothripsis, has received widespread attention since its discovery over a year ago. Until recently, chromothripsis was believed to originate from a single catastrophic event, but the molecular mechanisms leading to this event are yet to be uncovered. Because a thorough interpretation of the data are missing, the phenomenon itself has wrongly acquired the status of a mechanism used to justify many kinds of complex rearrangements. Although the assumption that all translocations in chromothripsis originate from a single event has met with criticism, satisfactory explanations for the intense but localized nature of this phenomenon are still missing. Here, we show why the data used to describe massive catastrophic rearrangements are incompatible with a model comprising a single event only and propose a molecular mechanism in which a combination of known cellular pathways accounts for chromothripsis. Instead of a single traumatic event, the protection of undamaged chromosomes by telomeres can limit repetitive breakage-fusion-bridge events to a single chromosome arm. Ultimately, common properties of chromosomal instability, such as aneuploidy and centromere fission, might establish the complex genetic pattern observed in this genomic state.     

Chromothripsis in Healthy Individuals Affects Multiple Protein-Coding Genes and Can Result in Severe Congenital Abnormalities in Offspring

Chromothripsis represents an extreme class of complex chromosome rearrangements (CCRs) with major effects on chromosomal architecture. Although recent studies have associated chromothripsis with congenital abnormalities, the incidence and pathogenic effects of this phenomenon require further investigation. Here, we analyzed the genomes of three families in which chromothripsis rearrangements were transmitted from a mother to her child. The chromothripsis in the mothers resulted in completely balanced rearrangements involving 8–23 breakpoint junctions across three to five chromosomes. Two mothers did not show any phenotypic abnormalities, although 3–13 protein-coding genes were affected by breakpoints. Unbalanced but stable transmission of a subset of the derivative chromosomes caused apparently de novo complex copy-number changes in two children. This resulted in gene-dosage changes, which are probably responsible for the severe congenital phenotypes of these two children. In contrast, the third child, who has a severe congenital disease, harbored all three chromothripsis chromosomes from his healthy mother, but one of the chromosomes acquired de novo rearrangements leading to copy-number changes. These results show that the human genome can tolerate extreme reshuffling of chromosomal architecture, including breakage of multiple protein-coding genes, without noticeable phenotypic effects. The presence of chromothripsis in healthy individuals affects reproduction and is expected to substantially increase the risk of miscarriages, abortions, and severe congenital disease.     

Chromothripsis is a common mechanism driving genomic rearrangements in primary and metastatic colorectal cancer

Background

Structural rearrangements form a major class of somatic variation in cancer genomes. Local chromosome shattering, termed chromothripsis, is a mechanism proposed to be the cause of clustered chromosomal rearrangements and was recently described to occur in a small percentage of tumors. The significance of these clusters for tumor development or metastatic spread is largely unclear.

Results

We used genome-wide long mate-pair sequencing and SNP array profiling to reveal that chromothripsis is a widespread phenomenon in primary colorectal cancer and metastases. We find large and small chromothripsis events in nearly every colorectal tumor sample and show that several breakpoints of chromothripsis clusters and isolated rearrangements affect cancer genes, including NOTCH2, EXO1 and MLL3. We complemented the structural variation studies by sequencing the coding regions of a cancer exome in all colorectal tumor samples and found somatic mutations in 24 genes, including APC, KRAS, SMAD4 and PIK3CA. A pairwise comparison of somatic variations in primary and metastatic samples indicated that many chromothripsis clusters, isolated rearrangements and point mutations are exclusively present in either the primary tumor or the metastasis and may affect cancer genes in a lesion-specific manner.

Conclusions

We conclude that chromothripsis is a prevalent mechanism driving structural rearrangements in colorectal cancer and show that a complex interplay between point mutations, simple copy number changes and chromothripsis events drive colorectal tumor development and metastasis.     

A cell‐based model system links chromothripsis with hyperploidy

A remarkable observation emerging from recent cancer genome analyses is the identification of chromothripsis as a one‐off genomic catastrophe, resulting in massive somatic DNA structural rearrangements (SRs). Largely due to lack of suitable model systems, the mechanistic basis of chromothripsis has remained elusive. We developed an integrative method termed “complex alterations after selection and transformation (CAST),” enabling efficient in vitro generation of complex DNA rearrangements including chromothripsis, using cell perturbations coupled with a strong selection barrier followed by massively parallel sequencing. We employed this methodology to characterize catastrophic SR formation processes, their temporal sequence, and their impact on gene expression and cell division. Our in vitro system uncovered a propensity of chromothripsis to occur in cells with damaged telomeres, and in particular in hyperploid cells. Analysis of primary medulloblastoma cancer genomes verified the link between hyperploidy and chromothripsis in vivo. CAST provides the foundation for mechanistic dissection of complex DNA rearrangement processes.     

Triggers for genomic rearrangements: insights into genomic, cellular and environmental influences

Genomic rearrangements are associated with many human genomic disorders, including cancers. It was previously thought that most genomic rearrangements formed randomly but emerging data suggest that many are nonrandom, cell type-, cell stage- and locus-specific events. Recent studies have revealed novel cellular mechanisms and environmental cues that influence genomic rearrangements. In this Review, we consider the multitude of influences on genomic rearrangements by grouping these influences into four categories: proximity of chromosomal regions in the nucleus, cellular stress, inappropriate DNA repair or recombination, and DNA sequence and chromatin features. The synergy of these triggers can poise a cell for rearrangements and here we aim to provide a conceptual framework for understanding the genesis of genomic rearrangements.     

Causes and consequences of micronuclei

Micronuclei are small membrane-bounded compartments with a DNA content encapsulated by a nuclear envelope and spatially separated from the primary nucleus. Micronuclei have long been linked to chromosome instability, genome rearrangements, and mutagenesis. They are frequently found in cancers, during senescence, and after genotoxic stress. Compromised integrity of the micronuclear envelope delays or disrupts DNA replication, inhibits DNA repair, and exposes micronuclear DNA directly to cytoplasm. Micronuclei play a central role in tumorigenesis, with micronuclear DNA being a source of complex genome rearrangements (including chromothripsis) and promoting a cyclic GMP–AMP synthase (cGAS)-mediated cellular immune response that may contribute to cancer metastasis. Here, we discuss recent findings on how micronuclei are generated, what the consequences are, and what cellular mechanisms can be applied to protect against micronucleation.     

Chromosome-breakage genomic instability and chromothripsis in breast cancer

Background

Chromosomal breakage followed by faulty DNA repair leads to gene amplifications and deletions in cancers. However, the mere assessment of the extent of genomic changes, amplifications and deletions may reduce the complexity of genomic data observed by array comparative genomic hybridization (array CGH). We present here a novel approach to array CGH data analysis, which focuses on putative breakpoints responsible for rearrangements within the genome.

Results

We performed array comparative genomic hybridization in 29 primary tumors from high risk patients with breast cancer. The specimens were flow sorted according to ploidy to increase tumor cell purity prior to array CGH. We describe the number of chromosomal breaks as well as the patterns of breaks on individual chromosomes in each tumor. There were differences in chromosomal breakage patterns between the 3 clinical subtypes of breast cancers, although the highest density of breaks occurred at chromosome 17 in all subtypes, suggesting a particular proclivity of this chromosome for breaks. We also observed chromothripsis affecting various chromosomes in 41% of high risk breast cancers.

Conclusions

Our results provide a new insight into the genomic complexity of breast cancer. Genomic instability dependent on chromosomal breakage events is not stochastic, targeting some chromosomes clearly more than others. We report a much higher percentage of chromothripsis than described previously in other cancers and this suggests that massive genomic rearrangements occurring in a single catastrophic event may shape many breast cancer genomes.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-579) contains supplementary material, which is available to authorized users.     

染色体粉碎——基因组灾难性事件产物

"染色体粉碎"是最初在肿瘤细胞中发现的一种复杂的基因组重排现象.在该事件中,细胞的一条或几条染色体在短时间内发生大量的DNA双链断裂,形成小的DNA碎片,之后这些碎片被细胞的DNA修复机制随机地拼接起来,形成新的染色体.染色体粉碎事件会造成大量的基因组重排,引起DNA拷贝数的变异和基因融合,从而导致正常细胞向肿瘤细胞的快速转化.这与传统的癌症发生理论不同,传统理论认为肿瘤的发生是由基因突变逐步积累而导致的,因此染色体粉碎现象可能揭示了一种肿瘤发生的新机制.目前,该现象的内在机制还不完全清楚,其判别标准也存在争议.本文综述了近年来关于染色体粉碎现象的判别标准和产生机制,探讨了该现象与肿瘤发生发展的关系,为进一步研究染色体粉碎事件提供参考.     

Chromothripsis and human disease: piecing together the shattering process

The unprecedented resolution of high-throughput genomics has enabled the recent discovery of a phenomenon by which specific regions of the genome are shattered and then stitched together via a single devastating event, referred to as chromothripsis. Potential mechanisms governing this process are now emerging, with implications for our understanding of the role of genomic rearrangements in development and disease.     

The hidden side of unstable DNA repeats: Mutagenesis at a distance

Structure-prone DNA repeats are common components of genomic DNA in all kingdoms of life. In humans, these repeats are linked to genomic instabilities that result in various hereditary disorders, including many cancers. It has long been known that DNA repeats are not only highly polymorphic in length but can also cause chromosomal fragility and stimulate gross chromosomal rearrangements, i.e., deletions, duplications, inversions, translocations and more complex shuffles. More recently, it has become clear that inherently unstable DNA repeats dramatically elevate mutation rates in surrounding DNA segments and that these mutations can occur up to ten kilobases away from the repetitive tract, a phenomenon we call repeat-induced mutagenesis (RIM). This review describes experimental data that led to the discovery and characterization of RIM and discusses the molecular mechanisms that could account for this phenomenon.     

Cancer cells are highly susceptible to accumulation of templated insertions linked to MMBIR

Microhomology-mediated break-induced replication (MMBIR) is a DNA repair pathway initiated by polymerase template switching at microhomology, which can produce templated insertions that initiate chromosomal rearrangements leading to neurological and metabolic diseases, and promote complex genomic rearrangements (CGRs) found in cancer. Yet, how often templated insertions accumulate from processes like MMBIR in genomes is poorly understood due to difficulty in directly identifying these events by whole genome sequencing (WGS). Here, by using our newly developed MMBSearch software, we directly detect such templated insertions (MMB-TIs) in human genomes and report substantial differences in frequency and complexity of MMB-TI events between normal and cancer cells. Through analysis of 71 cancer genomes from The Cancer Genome Atlas (TCGA), we observed that MMB-TIs readily accumulate de novo across several cancer types, with particularly high accumulation in some breast and lung cancers. By contrast, MMB-TIs appear only as germline variants in normal human fibroblast cells, and do not accumulate as de novo somatic mutations. Finally, we performed WGS on a lung adenocarcinoma patient case and confirmed MMB-TI-initiated chromosome fusions that disrupted potential tumor suppressors and induced chromothripsis-like CGRs. Based on our findings we propose that MMB-TIs represent a trigger for widespread genomic instability and tumor evolution.     

Starvation induces genomic rearrangements and starvation-resilient phenotypes in yeast

Evolution has shaped a wide variety of genomes across eukaryotic taxa. However, the forces that shape the genomes are generally unknown. Because organisms in nature commonly experience prolonged periods of nutrient depletion, we posit that diverse demographic, physiological, and genomic responses to starvation can occur. To test for these possibilities, we subjected replicate yeast populations to prolonged starvation. We observed that clones repeatedly gave rise to descendants that were karyotypically diverse. After a 1-month starvation period, approximately 70% of randomly isolated members of starved populations harbored one or more genomic rearrangements. Further, we found that 5 of 16 karyotypically differentiated groups of isolates from starved populations were more resilient to starvation than nonstarved clones and their common ancestor. Phylogenetic analysis of these isolates suggests that genomic rearrangements that arose during starvation can be adaptive in the context of a nutrient-depleted environment. Altogether our data illustrate the profound influence of environmental conditions on adaptive genome evolution in eukaryotes.     

Chromothripsis,a credible chromosomal mechanism in evolutionary process

Chromosoma - The recent discovery of a new class of massive chromosomal rearrangements, occurring during one unique cellular event and baptized “chromothripsis,” deeply modifies our...     

Cancer genomes evolve by pulverizing single chromosomes

A report in this issue describes "chromothripsis," a new mechanism for genetic instability in cancer cells. Chromothripsis appears to be a cataclysmic event in which a single chromosome is fragmented and then reassembled. The phenomenon raises important questions of how chromosome rearrangements can be confined to defined genome segments.     

Non-random distribution of instability-associated chromosomal rearrangement breakpoints in human lymphoblastoid cells

Genomic instability is observed in tumors and in a large fraction of the progeny surviving irradiation. One of the best-characterized phenotypic manifestations of genomic instability is delayed chromosome aberrations. Our working hypothesis for the current study was that if genomic instability is in part attributable to cis mechanisms, we should observe a non-random distribution of chromosomes or sites involved in instability-associated rearrangements, regardless of radiation quality, dose, or trans factor expression. We report here the karyotypic examination of 296 instability-associated chromosomal rearrangement breaksites (IACRB) from 118 unstable TK6 human B lymphoblast, and isogenic derivative, clones. When we tested whether IACRB were distributed across the chromosomes based on target size, a significant non-random distribution was evident (p < 0.00001), and three IACRB hotspots (chromosomes 11, 12, and 22) and one IACRB coldspot (chromosome 2) were identified. Statistical analysis at the chromosomal band-level identified four IACRB hotspots accounting for 20% of all instability-associated breaks, two of which account for over 14% of all IACRB. Further, analysis of independent clones provided evidence within 14 individual clones of IACRB clustering at the chromosomal band level, suggesting a predisposition for further breaks after an initial break at some chromosomal bands. All of these events, independently, or when taken together, were highly unlikely to have occurred by chance (p < 0.000001). These IACRB band-level cluster hotspots were observed independent of radiation quality, dose, or cellular p53 status. The non-random distribution of instability-associated chromosomal rearrangements described here significantly differs from the distribution that was observed in a first-division post-irradiation metaphase analysis (p = 0.0004). Taken together, these results suggest that genomic instability may be in part driven by chromosomal cis mechanisms.     

Genomic organization and evolution of double minutes/homogeneously staining regions with MYC amplification in human cancer

The mechanism for generating double minutes chromosomes (dmin) and homogeneously staining regions (hsr) in cancer is still poorly understood. Through an integrated approach combining next-generation sequencing, single nucleotide polymorphism array, fluorescent in situ hybridization and polymerase chain reaction-based techniques, we inferred the fine structure of MYC-containing dmin/hsr amplicons harboring sequences from several different chromosomes in seven tumor cell lines, and characterized an unprecedented number of hsr insertion sites. Local chromosome shattering involving a single-step catastrophic event (chromothripsis) was recently proposed to explain clustered chromosomal rearrangements and genomic amplifications in cancer. Our bioinformatics analyses based on the listed criteria to define chromothripsis led us to exclude it as the driving force underlying amplicon genesis in our samples. Instead, the finding of coexisting heterogeneous amplicons, differing in their complexity and chromosome content, in cell lines derived from the same tumor indicated the occurrence of a multi-step evolutionary process in the genesis of dmin/hsr. Our integrated approach allowed us to gather a complete view of the complex chromosome rearrangements occurring within MYC amplicons, suggesting that more than one model may be invoked to explain the origin of dmin/hsr in cancer. Finally, we identified PVT1 as a target of fusion events, confirming its role as breakpoint hotspot in MYC amplification.