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1.
Chromosome breakage and rejoining of sister chromatids in Bloom's syndrome   总被引:2,自引:0,他引:2  
The occurrence of chromosome breaks and reunion of sister chromatids in lymphocytes of two patients with Bloom's syndrome has been compared with those found in X-rayed and control cells. The distribution of breaks in BS is non-random both between and within chromosomes, the centric regions of certain chromosomes being preferentially involved. The following working hypotheses are put forward: When chromosome breaks in human lymphocytes occur in G0— G1, practically no sister chromatid reunion (SCR) takes place, whereas ends created by an S—G2 break show a considerable tendency to SCR. We propose further that chromosome aberrations in BS mainly result from breaks in S—G2, including possible U-type rejoining of sister chromatid exchanges. Fragments extra to an intact chromosome complement result from a chromatid break or an asymmetrical chromatid translocation in a previous mitosis.  相似文献   

2.
DNA double-strand breaks (DSBs) are arguably the most important lesions induced by ionizing radiation (IR) since unrepaired or misrepaired DSBs can lead to chromosomal aberrations and cell death. The two major pathways to repair IR-induced DSBs are non-homologous end-joining (NHEJ) and homologous recombination (HR). Perhaps surprisingly, NHEJ represents the predominant pathway in the G1 and G2 phases of the cell cycle, but HR also contributes and repairs a subset of IR-induced DSBs in G2. Following S-phase-dependent genotoxins, HR events give rise to sister chromatid exchanges (SCEs), which can be detected cytogenetically in mitosis. Here, we describe that HR occurring in G2-irradiated cells also generates SCEs in ∼50% of HR events. Since HR of IR-induced DSBs in G2 is a slow process, SCE formation in G2-irradiated cells requires several hours. During this time, irradiated S-phase cells can also reach mitosis, which has contributed to the widely held belief that SCEs form only during S phase. We describe procedures to measure SCEs exclusively in G2-irradiated cells and provide evidence that following IR cells do not need to progress through S phase in order to form SCEs.Key words: sister chromatid exchanges, double-strand break repair, ionizing radiation, homologous recombination, G2 phase  相似文献   

3.
Chinese hamster ovary cells (CHO) were X-irradiated in G1 and G2 stages of the cell cycle and subsequently Neurospora endonuclease (NE) (E.C.3.1.4), an enzyme which is specific in cleaving single-stranded DNA, was introduced into the cells, after making the cells permeable by treatment with inactivated Sendai virus. With this treatment all classes of X-ray-induced chromatid aberrations increased in G2 cells, whereas in G1 cells an increase in cromosome type of aberrations was found, associated with a profound induction of chromatid type of aberrations as well. Duration of the availability of single-strand gaps for the action of NE has been studied in G2 cells following X-irradiation and the influence of different parts of the G2 stage on the type and frequencies of chromatid aberrations was discerned. While the increase in chromosome type of aberrations by NE in X-irradiated G1 cells has been interpreted as due to the conversion of DNA single-strand breaks or gaps to double-strand breaks by NE, the induction of chromatid aberrations in G1 has been assumed to be due to conversion of some of the damaged bases strand breaks by NE. Biochemical evidence is presented for the conversion by NE of DNA single-strand breaks induced by X-rays into double-strand breaks using neutral sucrose gradient centrifugation.  相似文献   

4.
Three human malignancy cell lines were irradiated with 60Co γ-rays. Initial chromatid breaks were measured by using the chemically induced premature chromosome condensation technique. Survival curves of cells exposed to gamma rays was linear-quadratic while the efficiency of Calyculin A in inducing PCC of G2 PCC was about five times more than G1 PCC. A dose-dependent increase in radiation-induced chromatid/isochromatid breaks was observed in G1 and G2 phase PCC and a nearly positive linear correlation was found between cell survival and chromatin breaks. This study implies that low LET radiation-induced chromatid/isochromatid breaks can potentially be used to predict the radiosensitivity of tumor cells either in in vitro experimentation or in in vivo clinical radiotherapy.  相似文献   

5.
The effects of X-rays on the chromosomes of locust embryos   总被引:1,自引:0,他引:1  
D. P. Fox 《Chromosoma》1967,20(4):413-441
The variation in yield and the dose-response for chromatid aberration types following x-irradiation of Schistocerca gregaria embryo cells is described. Marked variations in yield are found for all aberration types during the G2 and latter part of S stages of interphase. Only gaps appear to follow similar curves, other aberration types having unique patterns of response. The dose exponents for the various chromatid aberration types are similar to, but lower than, those reported for other organisms. Chromatid “breaks” appear to have a dose exponent greater than 1.0 — a fact which is in conformation with the exchange hypothesis. The chromosome radiosensitivity of this organism is similar to that reported for other organisms.  相似文献   

6.
Seedlings of Crepis capillaris were irradiated after pulse-labelling with tritiated thymidine ([3H]TdR), and both chromosomal aberrations and presence of silver grains were recorded in the same metaphase cells at various intervals throughout the whole mitotic cycle. The following results were obtained: (a) irradiated roots were homogeneous with respect to the number of aberrations, and heterogenous with respect to labelling index (LI); (b) time-effect curves for labelled (L) and unlabelled (U) cells showed no significant difference from one another; (c) no significant quantitative difference of aberration spectra produced in S and G2 stages was found. These results support the view that the major factor which determines both quantitative and qualitative variation in the production of chromosomal aberrations by radiation is the time lapse between irradiation and fixation rather than relation of the time of irradiation to the time of DNA synthesis. In addition, it was found that labelling with [3H]TdR modifies the effect of radiation on chromosomes.  相似文献   

7.
S Wolff 《Mutation research》1972,15(4):435-444
The repair time for chromosome breaks induced by X-irradiation of unstimulated (G0) and stimulated (G1) human lymphocytes has been determined by dose fractionation studies. In both types of cells repair time was approx. 4–5 h. Treatment with hydroxyurea, a DNA synthesis inhibitor, did not prevent or delay the rejoining of broken chromosomes, whereas treatment with cycloheximide, a potent protein synthesis inhibitor, did. Thus, the repair of radiation-induced chromosome breaks in human lymphocytes is similar to the repair observed with plant cells.  相似文献   

8.
By means of combined experiments of X-irradiation and 3H-thymidine labeling of the chromosomes which are in the phase of synthesis, and the subsequent analysis at metaphase on the autoradiographs of the chromosomal damage induced during interphase, it was shown that in somatic cells from a quasi-diploid Chinese hamster line cultured in vitro the chromosomes change their response to radiation from single (chromosome type aberrations) to double (chromatid type aberrations) in late G1. These results are interpreted to indicate that the chromosome splits into two chromatids in G1, before DNA replication. — By extending the observations at the second metaphase after irradiation, it was also seen that cells irradiated while in G2 or late S when they reach the second post-irradiation mitosis still exhibit, beside chromosome type aberrations, many chromatid exchanges, some of which are labeled. Two hypotheses are suggested to account for this unexpected reappearance of chromatid aberrations at the second post-irradiation division. The first hypothesis is that they arise from half-chromatid aberrations. The second hypothesis, which derives from a new interpretation of the mechanisms of production of chromosome aberrations recently forwarded by Evans, is that they arise from gaps or achromatic lesions which undergo, as the cells go through the next cycle, a two-step repair process culminating in the production of aberrations.This work was supported in part by grant No. RH-00304 from the Division of Radiological Health, Bureau of State Services, Public Health Service, U.S.A.  相似文献   

9.
Cells in third mitosis treated during the first cell cycle with 3H-TdR and during the next two cycles with BrdU (without 3H-TdR) show a typical pattern of chromosome differentiation which allows identification of sister chromatid exchanges occurring during the first (SCE1, second (SCE2) and third cycles (SCE3). Chromosomes labeled only with 3H-TdR had the most SCEs; those labeled only with BrdU, the second highest number; and those labeled with 3H-TdR plus BrdU, the fewest. Since BrdU and 3H-TdR are well known inducers of SCEs, the relatively low frequency of exchanges produced by the combined action of these two compounds is paradoxical. — It is assumed that SCEs are generated by the abnormal recombination of double-strand DNA breaks occurring at the junctions between completely and partially duplicated replicon clusters. Thus, agents that induce absolute blocks to DNA fork displacement will favor the appearance of SCEs because double-strand breaks have more time to occur at junctions. Conversely, agents that inhibit the initiation of replication will decrease the probability of SCEs. Ionizing radiation delays the onset of cluster replication. Therefore, in 3H-TdR plus BrdU-substituted chromosomes the radiation from tritium may inhibit the appearance of BrdU-induced SCEs. Since the inhibition does not exist in chromosomes substituted only with BrdU, the frequency of SCEs in these elements is higher than in double-substituted chromosomes. During the first cell cycle the onset of cluster replication is normal. However, the incorporation of 3H-TdR in the replication fork may enhance the appearance of double-strand breaks, thus inducing a high frequency of SCEs.  相似文献   

10.
Synchronized G1 or G2 Chinese hamster cells were irradiated with UV light or X-rays and analyzed for chromosomal aberrations after one, two, or three replications. The cells were treated with Colcemid to induce polyploidy so that 2N, 4N, and 8N cells were scored. UV irradiation of G1 cells induces mainly chromatid aberrations, whereas X-rays induce chromosome aberrations. After both types of radiation chromatid aberrations appear in the polyploid cells. These results can be interpreted as indicating that UV and X-rays induce lesions at the subchromatid level that cannot be expressed until one or two replications have occurred. Since UV can induce long-lived lesions, the UV data do not allow us to choose between mononemic and polynemic models of the chromosome. X-rays, however, are ionizing radiations that might not produce long-lived lesions. The X-ray data, therefore, are more easily interpretable in terms of lesions being induced in the subunits of a polynemic chromosome.  相似文献   

11.
Wistar rats of both sexes were exposed to 100 R of X-rays. Chromatid-type aberrations in metaphase figures of bone marrow and lymph node cells were scored after 2, 4, 6, 8 and 24 h and 3, 5, 7, 9 and 24 h, respectively.The shape of the curve for chromatid plus isochromatid breaks in bone marrow cells versus time is exponential. It is suggested that this shape is mainly a consequence of the continuous entrance into mitosis of cells irradiated while in S phase, in addition to those that were irradiated in G2. For lymph nodes the frequency of chromatid plus isochromatid deletions increased up to the 5th h, then began to fall off in a manner similar to that for the bone marrow. The difference in the shape of the two curves is the consequence of the different dependence on time for chromatid and isochromatid breaks in each tissue. While the frequency of chromatid breaks fell steeply with time both for the bone marrow and for lymph nodes, the frequency of isochromatid breaks remained nearly constant for bone marrow, whereas it rose to a peak at the 5th h for the lymph nodes.These differences are tentatively explained by a shift in the phases of the cell cycle sampled owing to the greater mitotic delay of G2 cells in lymph nodes, with the suggestion that in the late S phase the frequency of isochromatid breaks is lower than in all other phases of the cell cycle.  相似文献   

12.
Effects of chlorambucil on human chromosomes   总被引:1,自引:0,他引:1  
No significant amount of chromosomal damage was found in the 48-h cultures of lymphocytes of 18 patients who had been treated with the bifunctional alkylating agent chlorambucil (CBC). However, there was suggestive evidence of chromatid damage (i.e. of types attributable to damage during or after DNA synthesis in the cell cycle). In marrow cells of 3 patients given a single large dose of chlorambucil (equivalent to 2 days' normal treatment) there was also suggestive evidence of induced chromatide-type damage.Extensive series of in vitro experiments yielded evidence that (a) exposure of human lymphocytes over the whole period of culture showed chromatid-type damage; (b) this damage increased sharply from concentrations of 0.5 μg/ml to3.0 μg/ml; (c) although chromatide-type damage always predominated, there was suggestive evidence also of chromosome-type aberrations attributable to damage occuring in the G0/G1 period, although some or all of this could be attributed to “derived” chromatid damage; (d) even if lymphocytes were only exposed during the G0 or G1 periods of the cycle, damage was found in the subsequent metaphases and it was almost entirely of the chromatid type; (e) much more damage occurred in lymphocytes exposed for varying periods to the drugs after stimulation by phytohaemagglutinins than in those exposed in whole blood, or in medium before stimulation; (f) damaged occurred in lymphocytes exposed to the drug while in S but not exposed only when in G2; (g) no evidence was found that unschaduled DNA synthesis during G0 or G1 was induced by the drug; (h) there appeared to be no delay caused by the drug in the time at which cells reached the first “S” phase in culture but there was some evidence consistent with prolongation of “S” in cells exposed in culture; (i) there was evidence that CBC alone could stimulate lymphocyte tto DNA synthesis, and that a few cells proceeded in the cycle to prophase, or even metaphase. However, there was a considerable amount of cell-killing during CBC-stimulated DNA synthesis.  相似文献   

13.
To increase the sensitivity of cytogenetic surveillance of exposure to mutagens in the peripheral lymphocyte assay, structural chromosome aberrations (CA) were studied after inhibition of DNA synthesis and DNA repair with hydroxyurea and caffeine in culture 3 h prior to harvesting. CA and sister-chromatid exchanges (SCE) from conventional cultures from the same subjects were used for comparison. Smoking was used as exposure parameter. Thirty-two smokers and 35 nonsmokers were studied. In the inhibited cultures a significantly higher number of aberrations was found in lymphocytes from smokers than nonsmokers: chromatid breaks (20.4 vs. 11.8, p = 0.0002), chromosome breaks (4.5 vs. 1.7, p = 0.0003), and the number of cells with aberrations (18.9 vs. 12.4, p = 0.0001), when 50 cells per subject were analyzed. In conventional cultures no increase in gaps, chromatid and chromosome breaks or number of cells with aberrations was found in smokers when 100 cells from each subject were studied. Smokers showed an increased number of SCE (6.8 vs. nonsmokers 5.9, p = 0.02). A significant positive linear correlation (r = 0.39, p = 0.01) was seen between SCE and the number of cells with chromatid breaks from inhibited cultures. The present results indicate that adding hydroxyurea and caffeine to lymphocyte cultures for the last 3 h prior to harvesting may enhance the detection of cytogenetic damage from previous in vivo exposure to mutagens.  相似文献   

14.
M. Gatti  C. Tanzarella    G. Olivieri 《Genetics》1974,77(4):701-719
A technique has been perfected for enabling good microscope preparations to be obtained from the larval ganglia of Drosophila melanogaster. This system was then tested with X-rays and an extensive series of data was obtained on the chromosome aberrations induced in the various stages of the cell cycle.-The analysis of the results obtained offers the following points of interest: (1) There exists a difference in radio-sensitivity between the two sexes. The females constantly display a greater frequency of both chromosome and chromatid aberrations. They also display a greater frequency of spontaneous aberrations. (2) In both sexes the overall chromosome damage is greater in cells irradiated in stages G(2) and G(1). These two peaks of greater radiosensitivity are produced by a high frequency of terminal deletions and chromatid exchanges and by a high frequency of dicentrics, respectively. (3) The aberrations are not distributed at random among the various chromosomes. On the average, the Y chromosome is found to be more resistant and the breaks are preferentially localized in the pericentromeric heterochromatin of the X chromosome and of the autosomes. (4) Somatic pairing influences the frequency and type of the chromosome aberrations induced. In this system, such an arrangement of the chromosomes results in a high frequency of exchanges and dicentrics between homologous chromosomes and a low frequency of scorable translocations. Moreover, somatic pairing, probably by preventing the formation of looped regions in the interphase chromosomes, results in the almost total absence of intrachanges at both chromosome and chromatid level.  相似文献   

15.
We have examined cell-cycle dependence of chromosomal aberration induction and cell killing after high or low dose-rate γ irradiation in cells bearing DNA-PKcs mutations in the S2056 cluster, the T2609 cluster, or the kinase domain. We also compared sister chromatid exchanges (SCE) production by very low fluences of α-particles in DNA-PKcs mutant cells, and in homologous recombination repair (HRR) mutant cells including Rad51C, Rad51D, and Fancg/xrcc9. Generally, chromosomal aberrations and cell killing by γ-rays were similarly affected by mutations in DNA-PKcs, and these mutant cells were more sensitive in G1 than in S/G2 phase. In G1-irradiated DNA-PKcs mutant cells, both chromosome- and chromatid-type breaks and exchanges were in excess than wild-type cells. For cells irradiated in late S/G2 phase, mutant cells showed very high yields of chromatid breaks compared to wild-type cells. Few exchanges were seen in DNA-PKcs-null, Ku80-null, or DNA-PKcs kinase dead mutants, but exchanges in excess were detected in the S2506 or T2609 cluster mutants. SCE induction by very low doses of α-particles is resulted from bystander effects in cells not traversed by α-particles. SCE seen in wild-type cells was completely abolished in Rad51C- or Rad51D-deficient cells, but near normal in Fancg/xrcc9 cells. In marked contrast, very high levels of SCEs were observed in DNA-PKcs-null, DNA-PKcs kinase-dead and Ku80-null mutants. SCE induction was also abolished in T2609 cluster mutant cells, but was only slightly reduced in the S2056 cluster mutant cells. Since both non-homologous end-joining (NHEJ) and HRR systems utilize initial DNA lesions as a substrate, these results suggest the possibility of a competitive interference phenomenon operating between NHEJ and at least the Rad51C/D components of HRR; the level of interaction between damaged DNA and a particular DNA-PK component may determine the level of interaction of such DNA with a relevant HRR component.  相似文献   

16.
p27kip1 is a cyclin-dependent kinase inhibitor and a tumor suppressor. In some tumors, p27 suppresses tumor growth by inhibition of cell proliferation. However, this is not universally observed, implying additional mechanisms of tumor suppression by p27. p27-deficient mice are particularly susceptibility to genotoxin-induced tumors, suggesting a role for p27 in the DNA damage response. To test this hypothesis, we measured genotoxin-induced mutations and chromosome damage in p27-deficient mice. Both p27+/− and p27−/− mice displayed a higher N-ethyl-N-nitrosourea-induced mutation frequency in the colon than p27+/+ littermates. Furthermore, cells from irradiated p27-deficient mice exhibited a higher number of chromatid breaks and showed modestly increased micronucleus formation compared to cells from wild-type littermates. To determine if this mutator phenotype was related to the cell cycle-inhibitory function of p27, we measured cell cycle arrest in response to DNA damage. Both normal and tumor cells from p27-deficient mice showed impaired G2/M arrest following low doses of ionizing radiation. Thus, p27 may inhibit tumor development through two mechanisms. The first is by reducing the proliferation of cells that have already sustained an oncogenic lesion. The second is by transient inhibition of cell cycle progression following genotoxic insult, thereby minimizing chromosome damage and fixation of mutations.  相似文献   

17.
Vicia faba root tip cells were treated for short periods with tritiated thymidine, either immediately before or after exposure of roots to x-rays, and autoradiograph preparations were analysed in an attempt to test the hypothesis that chromatid type (B') aberrations are induced only in those chromosome regions that have synthesized DNA prior to x-irradiation, whereas chromosome type (B') aberrations are induced only in unduplicated chromosome regions. Studying the relation between presence or absence of label at loci involved in aberrations, in cells irradiated at different development stages, and the pattern of labelling in cells carrying both types of aberration leads to the conclusion that B' aberrations are induced only in unreplicated chromosome regions. Following replication, only B' aberrations are induced, but these aberrations are also induced in chromosome regions preparing to incorporate DNA. It is suggested that the doubled response of the chromosome to x-rays prior to DNA incorporation might reflect a physical separation of replicating units prior to replication. The aberration yields in damaged cells which were irradiated in G1 S, and early G2 were in the ratio of 1.0:2.0:3.2. The data indicate that the increased yield of B' in early G2 relative to S cells may be a consequence of changes in the spatial distribution of the chromosomes within the nucleus.  相似文献   

18.
The efficient repair of double-strand breaks (DSBs) is crucial in maintaining genomic integrity. Sister chromatid cohesion is important for not only faithful chromosome segregation but also for proper DSB repair. During DSB repair, the Smc1–Smc3 cohesin complex is loaded onto chromatin around the DSB to support recombination-mediated DSB repair. In this study, we investigated whether Ctf18, a factor implicated in the establishment of sister chromatid cohesion, is involved in DSB repair in budding yeast. Ctf18 was recruited to HO-endonuclease induced DSB sites in an Mre11-dependent manner and to damaged chromatin in G2/M phase-arrested cells. The ctf18 mutant cells showed high sensitivity to DSB-inducible genotoxic agents and defects in DSB repair, as well as defects in damage-induced recombination between sister chromatids and between homologous chromosomes. These results suggest that Ctf18 is involved in damage-induced homologous recombination.  相似文献   

19.
Double-strand break (DSB) repair through homologous recombination (HR) is an evolutionarily conserved process that is generally error-free. The risk to genome stability posed by nonallelic recombination or loss-of-heterozygosity could be reduced by confining HR to sister chromatids, thereby preventing recombination between homologous chromosomes. Here we show that the sister chromatid cohesion complex (cohesin) is a limiting factor in the control of DSB repair and genome stability and that it suppresses DNA damage–induced interactions between homologues. We developed a gene dosage system in tetraploid yeast to address limitations on various essential components in DSB repair and HR. Unlike RAD50 and RAD51, which play a direct role in HR, a 4-fold reduction in the number of essential MCD1 sister chromatid cohesion subunit genes affected survival of gamma-irradiated G2/M cells. The decreased survival reflected a reduction in DSB repair. Importantly, HR between homologous chromosomes was strongly increased by ionizing radiation in G2/M cells with a single copy of MCD1 or SMC3 even at radiation doses where survival was high and DSB repair was efficient. The increased recombination also extended to nonlethal doses of UV, which did not induce DSBs. The DNA damage–induced recombinants in G2/M cells included crossovers. Thus, the cohesin complex has a dual role in protecting chromosome integrity: it promotes DSB repair and recombination between sister chromatids, and it suppresses damage-induced recombination between homologues. The effects of limited amounts of Mcd1and Smc3 indicate that small changes in cohesin levels may increase the risk of genome instability, which may lead to genetic diseases and cancer.  相似文献   

20.
The complementation effect of wild-type CHO-K1 and xrs mutants after fusion, as judged by the frequencies of X-ray-induced G1 and G2 premature chromosome condensation (PCC), was studied. For induction of PCC, X-irradiated interphase cells (G1 and G2) were fused immediately with untreated mitotic cells of the same cell line or with mitotic cells of another line. The frequencies of breaks in G1-PCC, or breaks and chromatid exchanges in G2-PCC were determined and the latter parameter was compared with the frequency of chromosomal aberrations in mitotic cells following G2 irradiation. CHO-K1 cells were capable of complementing the X-ray sensitivity of both xrs 5 and xrs 6 cells. However, full restoration of the repair defect in xrs cells could never be accomplished. The mutants failed to complement each other. In CHO-K1 cells, the incidence of chromosomal aberrations was significantly higher in G2-PCC (2.5-fold) than that observed in mitotic cells at 2.5 h after irradiation. The ratio of the induced frequency of aberrations in G2-PCC to that in mitotic cells was correlated with the degree of repair of DNA double-strand breaks (dsb) and reached almost 1 in xrs 5 cells indicating no repair. In addition the data indicated that, during the period of recovery of CHO-K1 cells, X-ray-induced breaks decreased but exchanges remained at the same level. In contrast, due to a deficiency in rejoining of dsb in xrs mutants, breaks remained open for a long period of time, allowing the formation of additional chromatid exchanges during recovery time.  相似文献   

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