Defining chromosomal translocation risks in cancer.
ABSTRACT: Chromosomal translocations are a hallmark of cancer. Unraveling the molecular mechanism of these rare genetic events requires a clear distinction between correlative and causative risk-determinants, where technical and analytical issues can be excluded. To meet this goal, we performed in-depth analyses of publicly available genome-wide datasets. In contrast to several recent reports, we demonstrate that chromosomal translocation risk is causally unrelated to promoter stalling (Spt5), transcriptional activity, or off-targeting activity of the activation-induced cytidine deaminase. Rather, an open chromatin configuration, which is not promoter-specific, explained the elevated translocation risk of promoter regions. Furthermore, the fact that gene size directly correlates with the translocation risk in mice and human cancers further demonstrated the general irrelevance of promoter-specific activities. Interestingly, a subset of translocations observed in cancer patients likely initiates from double-strand breaks induced by an access-independent process. Together, these unexpected and novel insights are fundamental in understanding the origin of chromosome translocations and, consequently, cancer.
Project description:Activation-induced cytidine deaminase (AID) initiates antibody gene diversification by creating U:G mismatches. However, AID is not specific for antibody genes; Off-target lesions can activate oncogenes or cause chromosome translocations. Despite its importance in these transactions little is known about how AID finds its targets. We performed an shRNA screen to identify factors required for class switch recombination (CSR) of antibody loci. We found that Spt5, a factor associated with stalled RNA polymerase II (Pol II) and single stranded DNA (ssDNA), is required for CSR. Spt5 interacts with AID, it facilitates association between AID and Pol II, and AID recruitment to its Ig and non-Ig targets. ChIP-seq experiments reveal that Spt5 colocalizes with AID and stalled Pol II. Further, Spt5 accumulation at sites of Pol II stalling is predictive of AID-induced mutation. We propose that AID is targeted to sites of Pol II stalling in part via its association with Spt5.
Project description:Gene rearrangements such as chromosomal translocations have been shown to contribute to cancer development. Human chromosomal fragile sites are regions of the genome especially prone to breakage, and have been implicated in various chromosome abnormalities found in cancer. However, there has been no comprehensive and quantitative examination of the location of fragile sites in relation to all chromosomal aberrations.Using up-to-date databases containing all cancer-specific recurrent translocations, we have examined 444 unique pairs of genes involved in these translocations to determine the correlation of translocation breakpoints and fragile sites in the gene pairs. We found that over half (52%) of translocation breakpoints in at least one gene of these gene pairs are mapped to fragile sites. Among these, we examined the DNA sequences within and flanking three randomly selected pairs of translocation-prone genes, and found that they exhibit characteristic features of fragile DNA, with frequent AT-rich flexibility islands and the potential of forming highly stable secondary structures.Our study is the first to examine gene pairs involved in all recurrent chromosomal translocations observed in tumor cells, and to correlate the location of more than half of breakpoints to positions of known fragile sites. These results provide strong evidence to support a causative role for fragile sites in the generation of cancer-specific chromosomal rearrangements.
Project description:Chromosomal translocations are rare in myelodysplastic syndrome (MDS) and their impact on overall survival (OS) and response to hypomethylating agents (HMA) is unknown. The prognostic impact of the revised International Prognostic Scoring System (IPSS-R) and for chromosomal translocations was assessed in 751 patients from the Korea MDS Registry. IPSS-R effectively discriminated patients according to leukaemia evolution risk and OS. We identified 40 patients (5.3%) carrying translocations, 30 (75%) of whom also fulfilled complex karyotype criteria. Translocation presence was associated with a shorter OS (median, 12.0 versus 79.7 months, P < 0.01). Multivariate analysis demonstrated that translocations (hazard ratio [HR] 1.64 [1.06-2.63]; P = 0.03) as well as age, sex, IPSS-R, and CK were independent predictors of OS. In the IPSS-R high and very high risk subgroup (n = 260), translocations remained independently associated with OS (HR 1.68 [1.06-2.69], P = 0.03) whereas HMA treatment was not associated with improved survival (median OS, 20.9 versus 21.2 months, P = 0.43). However, translocation carriers exhibited enhanced survival following HMA treatment (median 2.1 versus 12.4 months, P = 0.03). Our data suggest that chromosomal translocation is an independent predictor of adverse outcome and has an additional prognostic value in discriminating patients with MDS having higher risk IPSS-R who could benefit from HMA treatment.
Project description:For the largest class of human tumors, those of epithelial origin, little is known about their initiating genetic hits or cells of origin. Whether tissue stem cells or more committed progenitors are targets for transformation is also uncertain. Experience in hematopoietic malignancies and sarcomas teaches that recurrent chromosomal translocations represent initiating oncogenic events. To develop a system in which epithelial tumorigenesis can be assessed from the initial event to frank malignancy, we have generated mice that conditionally express the Etv6-NTRK3 (EN) fusion oncoprotein, the product of the t(12;15)(p13;q25) translocation characteristic of one form of human breast cancer. Activation of EN expression in mammary tissues by Whey acidic protein (Wap) promoter-driven Cre leads to fully penetrant, multifocal malignant breast cancer with short latency. We provide genetic evidence that committed, bipotent or CD61+ luminal alveolar progenitors, can be targets of tumorigenesis. Furthermore, EN transforms these otherwise transient progenitors through the AP1 complex. Our model supports the existence of an epithelial cell hierarchy in both normal mammary glands and malignancy. To our knowledge, this is the first murine model of human epithelial cancer based on a recurrent chromosomal translocation. Given increasing relevance of chromosomal translocations in epithelial cancers, such mice serve as a paradigm for the study of their genetic pathogenesis and cellular origins, and generation of novel preclinical models. Experiment Overall Design: Reference X Sample
Project description:Chromosomal translocations are frequent features of cancer genomes that contribute to disease progression. These rearrangements result from formation and illegitimate repair of DNA double-strand breaks (DSBs), a process that requires spatial colocalization of chromosomal breakpoints. The "contact first" hypothesis suggests that translocation partners colocalize in the nuclei of normal cells, prior to rearrangement. It is unclear, however, the extent to which spatial interactions based on three-dimensional genome architecture contribute to chromosomal rearrangements in human disease. Here we intersect Hi-C maps of three-dimensional chromosome conformation with collections of 1,533 chromosomal translocations from cancer and germline genomes. We show that many translocation-prone pairs of regions genome-wide, including the cancer translocation partners BCR-ABL and MYC-IGH, display elevated Hi-C contact frequencies in normal human cells. Considering tissue specificity, we find that translocation breakpoints reported in human hematologic malignancies have higher Hi-C contact frequencies in lymphoid cells than those reported in sarcomas and epithelial tumors. However, translocations from multiple tissue types show significant correlation with Hi-C contact frequencies, suggesting that both tissue-specific and universal features of chromatin structure contribute to chromosomal alterations. Our results demonstrate that three-dimensional genome architecture shapes the landscape of rearrangements directly observed in human disease and establish Hi-C as a key method for dissecting these effects.
Project description:The Myc-deregulating chromosomal T(12;15)(Igh-Myc) translocation, the hallmark mutation of inflammation- and interleukin 6-dependent mouse plasmacytoma (PCT), is the premier model of cancer-associated chromosomal translocations because it is the only translocation in mice that occurs spontaneously (B lymphocyte lineage) and with predictably high incidence (approximately 85% of PCT), and has a direct counterpart in humans: Burkitt lymphoma t(8;14)(q24;q32) translocation. Here, we report on the development of a genetic system for the detection of T(12;15)(Igh-Myc) translocations in plasma cells of a mouse strain in which an enhanced green fluorescent protein (GFP)-encoding reporter gene has been targeted to Myc. Four of the PCTs that developed in the newly generated translocation reporter mice, designated iGFP(5'Myc), expressed GFP consequent to naturally occurring T(12;15) translocation. GFP expression did not interfere with tumor development or the deregulation of Myc on derivative 12 of translocation, der (12), because the reporter gene was allocated to the reciprocal product of translocation, der (15). Although the described reporter gene approach requires refinement before T(12;15) translocations can be quantitatively detected in vivo, including in B lymphocyte lineage cells that have not yet completed malignant transformation, our findings provide proof of principle that reporter gene tagging of oncogenes in gene-targeted mice can be used to elucidate unresolved questions on the occurrence, distribution and trafficking of cells that have acquired cancer-causing chromosomal translocations of great relevance for humans.
Project description:Recurrent reciprocal translocations are present in many hematologic and mesenchymal malignancies. Because significant sequence homology is absent from translocation breakpoint junctions, non-homologous end-joining (NHEJ) pathways of DNA repair are presumed to catalyze their formation. We developed translocation reporters for use in mammalian cells from which NHEJ events can be selected after precise chromosomal breakage. Translocations were efficiently recovered with these reporters using mouse cells, and their breakpoint junctions recapitulated findings from oncogenic translocations. Small deletions and microhomology were present in most junctions; insertions and more complex events also were observed. Thus, our reporters model features of oncogenic rearrangements in human cancer cells. A homologous sequence at a distance from the break site affected the translocation junction without substantially altering translocation frequency. Interestingly, in a direct comparison, the spectrum of translocation breakpoint junctions differed from junctions derived from repair at a single chromosomal break, providing mechanistic insight into translocation formation.
Project description:Chromosomal translocations result from joining of DNA double-strand breaks (DSBs) and frequently cause cancer. Yet, the steps linking DSB formation to DSB ligation remain undeciphered. We report that DNA replication timing (RT) directly regulates lymphomagenic Myc translocations during antibody maturation in B-cells downstream of DSBs and independently of DSB frequency. Depletion of minichromosome-maintenance (MCM) complexes alters replication origin activity, decreases translocations and abrogates global RT. Ablating a single origin at Myc causes an early-to-late RT switch, loss of translocations and reduced nuclear proximity with a translocation partner locus, phenotypes that were reversed by restoring early RT. Disruption of shared early RT also reduced tumorigenic translocations in human leukemic cells. Thus, RT constitutes a new, unprecedented mechanism in translocation biogenesis linking DSB formation to DSB ligation
Project description:Genome instability, epigenetic remodelling and structural chromosomal rearrangements are hallmarks of cancer. However, the coordinated epigenetic effects of constitutional chromosomal rearrangements that disrupt genes associated with congenital neurodevelopmental diseases are poorly understood. To understand the genetic-epigenetic interplay at breakpoints of chromosomal translocations disrupting CG-rich loci, we quantified epigenetic modifications at DLGAP4 (SAPAP4), a key post-synaptic density 95 (PSD95) associated gene, truncated by the chromosome translocation t(8;20)(p12;q11.23), co-segregating with cerebellar ataxia in a five-generation family. We report significant epigenetic remodelling of the DLGAP4 locus triggered by the t(8;20)(p12;q11.23) translocation and leading to dysregulation of DLGAP4 expression in affected carriers. Disruption of DLGAP4 results in monoallelic hypermethylation of the truncated DLGAP4 promoter CpG island. This induced hypermethylation is maintained in somatic cells of carriers across several generations in a t(8;20) dependent-manner however, is erased in the germ cells of the translocation carriers. Subsequently, chromatin remodelling of the locus-perturbed monoallelic expression of DLGAP4 mRNAs and non-coding RNAs in haploid cells having the translocation. Our results provide new mechanistic insight into the way a balanced chromosomal rearrangement associated with a neurodevelopmental disorder perturbs allele-specific epigenetic mechanisms at breakpoints leading to the deregulation of the truncated locus.