SEARCH · Search PubMed
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Cellular metabolic rates might regulate aging by impinging on genomic stability through the DNA repair pathways. A new study published in Cell (Mostoslavsky et al., 2006) reports that deficiency in one of the mammalian Sir2 homologs, SIRT6, results in genome instability through the DNA base excision repair pathway and leads to aging-associated degenerative phenotypes.
Genomic instability promotes tumorigenesis and can occur through various mechanisms, including defective segregation of chromosomes or inactivation of DNA mismatch repair. Although B-cell lymphomas are associated with chromosomal translocations that deregulate oncogene expression, a mechanism for genome-wide instability during lymphomagenesis has not been described. During B-cell development, the immunoglobulin variable (V) region genes are subject to somatic hypermutation in germinal-centre B cells. Here we report that an aberrant hypermutation activity targets multiple loci, including the proto-oncogenes PIM1, MYC, RhoH/TTF (ARHH) and PAX5, in more than 50% of diffuse large-cell lymphomas (DLCLs), which are tumours derived from germinal centres. Mutations are distributed in the 5' untranslated or coding sequences, are independent of chromosomal translocations, and share features typical of V-region-associated somatic hypermutation. In contrast to mutations in V regions, however, these mutations are not detectable in normal germinal-centre B cells or in other germinal-centre-derived lymphomas, suggesting a DLCL-associated malfunction of somatic hypermutation. Intriguingly, the four hypermutable genes are susceptible to chromosomal translocations in the same region, consistent with a role for hypermutation in generating translocations by DNA double-strand breaks. By mutating multiple genes, and possibly by favouring chromosomal translocations, aberrant hypermutation may represent the major contributor to lymphomagenesis.
A number of ongoing delayed effects have now been described in the progeny of an irradiated cell. These are grouped under the rubric of radiation induced genomic instability. Perhaps the best characterized is the dynamic production of chromosomal rearrangements in some clonally expanded cells surviving irradiation. In this chapter we provide the protocols for irradiation, cell culture, chromosome analysis, and characterization of the status of genomic stability in the context of delayed radiation effects.
Repetitive sequences constitute landmarks for genome regulation, evolution, and chromatin architecture. Patterns of specific and non-specific repetitive sequences change in many types and stages of tumor cells, characterized by band loss, gain, and (de) increased staining of pre-existing bands. In this work, repetitive DNA was studied in search of genome instability of skin cancers: basal and squamous cell carcinomas (BCC and SCC), malignant melanoma (MM), melanocytic nevus (MN), and actinic keratosis (AK) lesions. DNAs were extracted from blood and tumor samples from 21 BCC, 7 SCC, 11 MM and 7 lesions. Banding patterns were obtained by random amplification of polymorphic DNA (RAPD), and specific D9S50 and D9S52 microsatellites (9p21). D9S50 patterns revealed microsatellite instability (MSI) and/or loss of heterozygosity (LOH) in 36% BCC, 25% SCC, and 57% MM tumors. D9S52 microsatellite showed 28.5%; 42.8%; and 71.4% altered tumors, respectively. No microsatellite alterations were found in MN and AK. On the other hand, genomic rearrangements detected by RAPD were present in 100% tumors. In BCC, the mean number of tumor DNA alterations showed predominant gain of bands. On the contrary, MM samples presented loss, or decreased intensity signal of RAPD bands. Genome alterations in skin cancers would result from chromosomal rearrangements, aneuploidy and/or polysomies. The low-cost and quick RAPD technique may reveal unknown genes or DNA sequences associated with tumor development and progression, and may be easily implemented in clinical diagnosis.
Poly(ADP-ribose)polymerase 1 (PARP1) is well characterized for its role in base excision repair (BER), where it is activated by and binds to DNA breaks and catalyzes the poly(ADP-ribosyl)ation of several substrates involved in DNA damage repair. Here we demonstrate that PARP1 associates with telomere repeat binding factor 2 (TRF2) and is capable of poly(ADP-ribosyl)ation of TRF2, which affects binding of TRF2 to telomeric DNA. Immunostaining of interphase cells or metaphase spreads shows that PARP1 is detected sporadically at normal telomeres, but it appears preferentially at eroded telomeres caused by telomerase deficiency or damaged telomeres induced by DNA-damaging reagents. Although PARP1 is dispensable in the capping of normal telomeres, Parp1 deficiency leads to an increase in chromosome end-to-end fusions or chromosome ends without detectable telomeric DNA in primary murine cells after induction of DNA damage. Our results suggest that upon DNA damage, PARP1 is recruited to damaged telomeres, where it can help protect telomeres against chromosome end-to-end fusions and genomic instability.
The chromosomally integrated shuttle vector pUR288 contains a lacZ reporter gene to study mutagenesis in vivo. We used pUR288 to compare patterns of genomic instability in two mouse models, lymphoma resulting from deregulated c-MYC expression (lambda-MYC), and endogenous oxidative stress caused by partial glucose 6-phosphate dehydrogenase (G6PD) deficiency. We found previously that spontaneous mutations in both models were predominantly genomic rearrangements of lacZ with mouse sequences, while most mutations in controls were point mutations. Here, we characterized the fine structure of 68 lacZ/mouse rearrangements from lambda-MYC lymphomas and G6PD deficient mice by sequencing breakpoint junctions and determining the origin of recombining mouse sequences. Fifty-eight of 68 (85%) recombination partners were identified. The structure of rearrangements from both lambda-MYC and G6PD deficient mice were remarkably alike. Intra-chromosomal deletions and inversions were common, occurring in 41% (24/58) of rearrangements, while 59% (34/58) were random translocations between lacZ and other chromosomes. Signatures of double strand break repair by nonhomologous end joining were observed at breakpoint junctions; 37% (25/68) contained 1-4 bp microhomologies, while the remaining breakpoints had no sequence homology. Long interspersed nuclear element-1 (LINE-1 or L1) retrotransposons, which constitute approximately 10% of the mouse genome, were present at 25% (17/68) of breakpoints, suggesting their participation in rearrangements. The similarity in the structure of rearrangements is consistent with the hypothesis that genetic rearrangements in lambda-MYC lymphomas and G6PD deficient mice result from the same mechanism, mutagenic repair of DNA double strand breaks arising from oxidative damage.
Certain chromosome rearrangements, found in cancer cells or in cells exposed to ionizing radiation, exhibit a chromosome-wide delay in replication timing (DRT) that is associated with a delay in mitotic chromosome condensation (DMC). We have developed a chromosome engineering strategy that allows the generation of chromosomes with this DRT/DMC phenotype. We found that approximately 10% of inter-chromosomal translocations induced by two distinct mechanisms, site-specific recombination mediated by Cre or non-homologous end joining of DNA double-strand breaks induced by I-Sce1, result in DRT/DMC. Furthermore, on certain balanced translocations only one of the derivative chromosomes displays the phenotype. Finally, we show that the engineered DRT/DMC chromosomes acquire gross chromosomal rearrangements at an increased rate when compared with non-DRT/DMC chromosomes. These results indicate that the DRT/DMC phenotype is not the result of a stochastic process that could occur at any translocation breakpoint or as an epigenetic response to chromosome damage. Instead, our data indicate that the replication timing of certain derivative chromosomes is regulated by a cis-acting mechanism that delays both initiation and completion of DNA synthesis along the entire length of the chromosome. Because chromosomes with DRT/DMC are common in tumor cells and in cells exposed to ionizing radiation, we propose that DRT/DMC represents a common mechanism responsible for the genomic instability found in cancer cells and for the persistent chromosomal instability associated with cells exposed to ionizing radiation.
Cancers induced by ionizing radiation have no particular specificity nor genetic remarkable signature, excepting numerous multideletions. They should therefore be studied in the general field of cancer biology in its broad sense. A gap remains between the initial events like the rather well identified genomic damage and the subsequent emerging cellular clone with cancer characteristics. Intermediate steps are generally described as accumulation of mutations and epigenetic modifications leading at one point to the malignant phenotype. However we have no clear nor understandable model on these steps of malignant transformation till now. It is quite possible that specific causes (tobacco, alcohol, radiations, chemical toxics) which produce different initial abnormalities then lead to (or accelerate the entry in) the common and same way as that resulting of accumulations of damage due to ageing. Genomic instability is certainly an important factor involved in the cellular drift leading to malignant transformation. We postulate that only cells having both a high telomerase activity and a low apoptotic activity may become cancerous. The hypothesis is that cancer results from a genome reprogramming of these cells due to an oncogenic mitotic pressure which induces a loss of the differentiation control. We propose to name anti-apoptosis, in contrast to apoptosis which is the programmed death, the ultimate process by which a cell loses its tissue-related properties. The oncogenic activation may propagate to primordial genes of development resulting in emergence of a subnuclear with tumoral activity. Bio-molecular studies of embryonic development and of genome re-programming will probably allow us to better understand the mechanisms of cancer.
Proliferating cell nuclear antigen (PCNA), a sliding clamp required for processive DNA synthesis, provides attachment sites for various other proteins that function in DNA replication, DNA repair, cell cycle progression and chromatin assembly. It has been shown that differential posttranslational modifications of PCNA by ubiquitin or SUMO play a pivotal role in controlling the choice of pathway for rescuing stalled replication forks. Here, we explored the roles of Mgs1 and PCNA in replication fork rescue. We provide evidence that Mgs1 physically associates with PCNA and that Mgs1 helps suppress the RAD6 DNA damage tolerance pathway in the absence of exogenous DNA damage. We also show that PCNA sumoylation inhibits the growth of mgs1 rad18 double mutants, in which PCNA sumoylation and the Srs2 DNA helicase coordinately prevent RAD52-dependent homologous recombination. The proposed roles for Mgs1, Srs2, and modified PCNA during replication arrest highlight the importance of modulating the RAD6 and RAD52 pathways to avoid genome instability.
The breast/ovarian cancer susceptibility proteins BRCA1 and BRCA2 maintain genome stability, at least in part, through a functional role in DNA damage repair. They both colocalize with RAD51 at sites of DNA damage/replication and activate RAD51-mediated homologous recombination repair of DNA double-strand breaks (DSB). Whereas BRCA2 interacts directly with and regulates RAD51, the role of BRCA1 in this process is unclear. However, BRCA1 may regulate RAD51 in response to DNA damage or through its ability to interact with and regulate MRE11/RAD50/NBS1 (MRN) during the processing of DSBs into single-strand DNA (ssDNA) ends, prerequisite substrates for RAD51, or both. To test these hypotheses, we measured the effect of BRCA1 on the competition between RAD51-mediated homologous recombination (gene conversion and crossover) versus RAD51-independent homologous recombination [single-strand annealing (SSA)] for ssDNA at a site-specific chromosomal DSB within a DNA repeat, a substrate for both homologous recombination pathways. Expression of wild-type BRCA1 in BRCA1-deficient human recombination reporter cell lines promoted both gene conversion and SSA but greatly enhanced gene conversion. In addition, BRCA1 also suppressed both spontaneous gene conversion and deletion events, which can arise from either crossover or sister chromatid replication slippage (SCRS), a RAD51-independent process. BRCA1 does not seem to block crossover. From these results, we conclude that (a) BRCA1 regulates RAD51 function in response to the type of DNA damage and (b) BRCA1 suppresses SCRS, suggesting a role for this protein in sister chromatid cohesion/alignment. Loss of such control in response to estrogen-induced DNA damage after BRCA1 inactivation may be a key initial event that triggers genome instability and carcinogenesis.
The aim of the research is a further analysis of a problem concerning two (regulatory and damaging) functions of reactive oxygen species (ROS) in viability of organism cells under acute exposure to ionizing radiation. For this purpose the ROS content and the state of DNA structure in bone marrow cells of male CBA and SHK mice have been studied in dynamics, from 15 minutes up to 185 day after acute exposure to a sublethal dose (1.5 Gy) of ionizing radiation. The analysis of dependencies between these parameters in the norm, immediately after irradiation and in later cell descendants showed the direct correlation between the ROS content and the DNA nativity in the norm; 185 days after irradiation the correlation disappeared. It was suggested that the correlation occurred in the norm indicates participation of the ROS (as a sensory link) in a system of reactions (under the control of the corresponding genetic program), that ensure the DNA structure and, ultimately, the genome stability. The loss of such connection after acute exposure to ionizing radiation indicates actuation of another module of reactions sustaining stability of cellular genome in new conditions, without regulatory participation of ROS, that can promote or demonstrate the development of radiation-induced genome instability.
Of the various types of DNA damage that can occur within the mammalian cell, the DNA double strand break (DSB) is perhaps the most dangerous. DSBs are typically induced by intrinsic sources such as the by products of cellular metabolism or by extrinsic sources such as X-rays or gamma-rays and chemotherapeutic drugs. It is becoming increasing clear that an inability to respond properly to DSBs will lead to genomic instability and promote carcinogenesis. The mammalian cell, therefore, has in place several mechanisms that can respond rapidly to DSBs. In this review, we focus on the role of one such mechanism, the non-homologous end joining (NHEJ) pathway of DSB repair, in maintaining genome integrity and preventing carcinogenesis.
Chromosomal translocations and genomic instability are universal hallmarks of tumor cells. While the molecular mechanisms leading to the formation of translocations are rapidly being elucidated, a cell biological understanding of how chromosomes undergo translocations in the context of the cell nucleus in vivo is largely lacking. The recent realization that genomes are non-randomly arranged within the nuclear space has profound consequences for mechanisms of chromosome translocations. We review here the emerging principles of spatial genome organization and discuss the implications of non-random spatial genome organization for the genesis and specificity of cancerous chromosomal translocations.
Among the cellular responses observed following treatment with DNA-damaging agents is the activation of Short Interspersed Elements (SINEs; retrotransposable genetic elements that comprise over 10% of the human genome). By placing a human SINE (the Alu element) into murine cells, we have previously shown that DNA-damaging agents such as etoposide can induce both upregulation of SINE transcript levels and SINE retrotransposition. A similarly cytotoxic (but not genotoxic) exposure to vincristine was not associated with SINE activation. Here we demonstrate that multiple other genotoxic exposures are associated with upregulation of SINE transcript levels. By comparing the effects of similarly cytotoxic doses of the topoisomerase II inhibitors etoposide and merbarone, we confirm that DNA strand breakage is specifically associated with SINE induction. By evaluating transcription rate and RNA stability, we demonstrate that SINE induction by genotoxic exposure is associated with transcriptional induction and not with transcript stabilization. Finally we demonstrate that SINE induction by genotoxic stress is mediated by a Trp53-independent pathway, and in fact that Trp53 plays an inhibitory role in attenuating the transcriptional induction of SINE elements following exposure to a genotoxic agent. Together these data support a model in which initial DNA damage can trigger genomic instability due to SINE activation, a response which may be amplified in cancer cells lacking functional TP53.
Simple sequence repeats (microsatellites) are found in all eukaryotic genomes. Instabilities within these sequences have been associated with several human disorders including Huntington's chorea and myotonic dystrophy. Further studies have identified links between microsatellite instability, faulty mismatch repair and certain human cancers, in particular a form of hereditary colorectal cancer. The assay system described here consists of a congenic set of yeast strains mutated in DNA replication and mismatch repair genes and assay plasmids with which it is possible to measure differences in microsatellite stability in the range of 5-850-fold. The development of this technology will allow monitoring of environmental and dietary influences on the genomic stability in the context of human disease.
Down syndrome (DS) is a multifactorial disorder with a high predisposition to leukemia and other malignancies. A change in the replication pattern from synchronous in normal genes to asynchronous in DS amniocytes has previously been reported. The objective of this study was to evaluate additional molecular cytogenetic factors which could re-emphasize the high correlation between DS cells and genetic instability. We found a higher rate of random aneuploidy in chromosomes 9 and 18 and a higher rate of asynchronous replication in the subtelomeric region or DS leukocytes than in cells from normal newborns. In addition, the telomere capture phenomenon was observed in the DS leukocytes but not in normal controls. The molecular cytogenetic factors observed in the DS individuals are known to correlate with genomic instability and with predisposition to cancer.
Radiation-induced lymphomagenesis and leukemogenesis are complex processes involving both genetic and epigenetic changes. Although genetic alterations during radiation-induced lymphoma- and leukemogenesis are fairly well studied, the role of epigenetic changes has been largely overlooked. Rodent models are valuable tools for identifying molecular mechanisms of lymphoma and leukemogenesis. A widely used mouse model of radiation-induced thymic lymphoma is characterized by a lengthy "pre-lymphoma" period. Delineating molecular changes occurring during the pre-lymphoma period is crucial for understanding the mechanisms of radiation-induced leukemia/lymphoma development. In the present study, we investigated the role of radiation-induced DNA methylation changes in the radiation carcinogenesis target organ--thymus, and non-target organ--muscle. This study is the first report on the radiation-induced epigenetic changes in radiation-target murine thymus during the pre-lymphoma period. We have demonstrated that acute and fractionated whole-body irradiation significantly altered DNA methylation pattern in murine thymus leading to a massive loss of global DNA methylation. We have also observed that irradiation led to increased levels of DNA strand breaks 6 h following the initial exposure. The majority of radiation-induced DNA strand breaks were repaired 1 month after exposure. DNA methylation changes, though, were persistent and significant radiation-induced DNA hypomethylation was observed in thymus 1 month after exposure. In sharp contrast to thymus, no significant persistent changes were noted in the non-target muscle tissue. The presence of stable DNA hypomethylation in the radiation-target tissue, even though DNA damage resulting from initial genotoxic radiation insult was repaired, suggests of the importance of epigenetic mechanisms in the development of radiation-related pathologies. The possible role of radiation-induced DNA hypomethylation in radiation-induced genome instability and aberrant gene expression in molecular etiology of thymic lymphomas is discussed.
The Drosophila melanogaster warts/lats tumour suppressor has two mammalian counterparts LATS1/Warts-1 and LATS2/Kpm. Here, we show that mammalian Lats orthologues exhibit distinct expression profiles according to germ cell layer origin. Lats2(-/-) embryos show overgrowth in restricted tissues of mesodermal lineage; however, lethality ultimately ensues on or before embryonic day 12.5 preceded by defective proliferation. Lats2(-/-) mouse embryonic fibroblasts (MEFs) acquire growth advantages and display a profound defect in contact inhibition of growth, yet exhibit defective cytokinesis. Lats2(-/-) embryos and MEFs display centrosome amplification and genomic instability. Lats2 localizes to centrosomes and overexpression of Lats2 suppresses centrosome overduplication induced in wild-type MEFs and reverses centrosome amplification inherent in Lats2(-/-) MEFs. These findings indicate an essential role of Lats2 in the integrity of processes that govern centrosome duplication, maintenance of mitotic fidelity and genomic stability.