共查询到20条相似文献,搜索用时 31 毫秒
1.
Naoto Ohnishi Hiroaki Kodania Satoshi Ando Atsushi Komamine 《Physiologia plantarum》1990,80(1):95-101
The effects of inhibition of the synthesis of protein, mRNA or rRNA on the progression of the cell cycle have been analyzed in cultures of Catharanthus roseus in which cells were induced to divide in synchrony by the double phosphate starvation method. The partial inhibition of protein synthesis at the G1 phase by anisoniycio or cycloheximide caused the arrest of cells in the G1 phase or delayed the entry of cells into the S phase. When protein synthesis was partially inhibited at the S phase, cell division occurred to about the same extent as in the control. When asynchronously dividing cells were treated with cycloheximide, cells accumulated in the G1 phase, as shown by flow-cytometric analysis. The partial inhibition of mRNA synthesis by α-amanitin at the G1 phase caused the arrest of cells in the G1 phase, although partial inhibition of mRNA synthesis at the S phase had little effect on cell division. In the case of inhibition of synthesis of rRNA by actinomycin D at the G1 phase, initiation of DNA synthesis was observed, but no subsequent DNA synthesis or the division of cells occurred. However, the addition of actinomycin D during the S phase had no effect on cell division. These results suggest that specific protein(s), required for the progression of the cell cycle, are synthesized in the G1 phase, and that the mRNA(s) that encode these proteins are also synthesized at the G1 phase. 相似文献
2.
Actinomycin D (0.5 μg/ml) did not prevent M stage cells from entering G1 stage, but blocked their progress from G1 to S stage. The position of the block was approximately 1.4 hr before S stage or just after the beginning of G1 stage. Actinomycin D in this concentration also significantly depressed uridine-3 H uptake into G1 stage cells, but did not suppress leucine-3 H uptake by M and G1 cells. This suggests that some proteins may be synthesized in M and G1 stage cells by messenger RNA left over from the previous cell cycle. However, entry of G1 cells into S stage would require synthesis of new messenger RNA near the beginning of G1 stage. Puromycin (10 μg/ml) did not prevent M cells from entering G1 stage, but blocked their progress from G1 to S stage. The site of blockage was about 0.7 hr before S stage or in the first two-third of G1 stage. This might be the site where the cells synthesize new G1 proteins necessary for entry to S stage.
Comparison of sensitivities of G1 and G2 stages to the two antibiotics reveals that the puromycin sensitivity of G1 cells was similar to that of G2 cells, but the actinomycin D sensitivity of G1 was greater than that of G2 cells. 相似文献
Comparison of sensitivities of G
3.
Cell cycle progression in human cells following re-oxygenation after extreme hypoxia: consequences concerning initiation of DNA synthesis 总被引:1,自引:0,他引:1
Abstract. The initiation of DNA synthesis and further cell cycle progression in cells during and following exposure to extremely hypoxic conditions in either G1 or G2 +M has been studied in human NHIK 3025 cells. Populations of cells, synchronized by mitotic selection, were rendered extremely hypoxic (< 4 p.p.m. O2 ) for up to 24n h. Cell cycle progression was studied from flow cytometric DNA recordings. No accumulation of DNA was found to take place during extreme hypoxia. Cells initially in G1 at the onset of treatment did not enter S during up to 24 h exposure to extreme hypoxia, but started DNA synthesis in a highly synchronous manner within 1.5 to 2.25 h after reoxygenation. The duration of S phase was only slightly affected (increased by ≅10%) by the hypoxic treatment. This suggests that the DNA synthesizing machinery either remains intact during hypoxia or is rapidly restored after reoxygenation. Cells initially in G2 at the onset of hypoxia were able to complete mitosis, but further cell cycle progression was blocked in the subsequent G^ Following reoxygenation, these cells progressed into S phase, but the initiation of DNA synthesis was delayed for a period corresponding to at least the duration of normal G1 and did not appear in a synchronous manner. In fact, cell cycle variability was found to be increased rather than decreased as a result of exposure to hypoxia starting in G2 . We interpret these findings as an indication that important steps in the preparation for initiation of DNA synthesis take place before mitosis. Furthermore, the change in cell cycle duration induced by hypoxia commencing in G1 is of a nature other than that induced by hypoxia commencing in other parts of the cell cycle. 相似文献
4.
Abstract. We have previously found that DNA replication was affected within one cell cycle after seeding Chinese hamster ovary (CHO) cells in the presence of the polyamine biosynthesis inhibitor 2-difluoromethylornithine (DFMO). We could, however, not rule out if this was due to an effect on the G1 /S transition and/or on DNA synthesis elongation. In the present paper, we use a bromodeoxyuridine-flow cytometric method to more specifically study the G1 /S transition, the S phase length, and the progression of cells from S phase through G2 + M and into G1 , after seeding plateau phase CHO cells at low density in the absence or presence of 5 mM DFMO. We report here that DFMO-induced polyamine depletion increased the length of the S phase within one cell cycle after seeding of CHO cells in the presence of the inhibitor. No effect on the G1 /S transition was observed until 2 days after seeding, suggesting that a DFMO-induced lengthening of the G1 phase occurred later than the effect on S phase progression. These results imply that the G2 + M phase was not prolonged until 2 days after seeding CHO cells in the presence of DFMO. 相似文献
5.
D. L. Dewey 《Cell proliferation》1987,20(6):603-609
Abstract When cells in culture are released from G0 into cycle by diluting into fresh medium there is a delay of many hours before they re-enter the cycle and start DNA synthesis. A mouse melanoma cell line designated HP2 has been used to investigate the effects of non-standard temperatures between the time of plating and DNA synthesis. When the cells were incubated in a 5% CO2 box at 8° C for periods during the G0 -G1 transition there was an extra delay before the start of S, approximately equal to the time that the cells were held at 8° C and independent of the time when the cold pulse was administered. When the cells were cooled to 25° C the delay was longer than the time for which the cells had been kept at 25° C, and this extra delay was also dependent on the point in G0 -G1 when the cells were cooled, as though the cells could be reset to an earlier time by this treatment. It is suggested that a labile substance required for progression is destroyed faster than it is made at 25° C but at 8° C the rate of destruction is very low. Another phenomenon noted during these cooling experiments was that the peak height of the S phase profile, as measured by frequent pulse-thymidine incorporation experiments, was substantially higher for cells which had been cooled at a later stage in the G0 -G1 transition, even though the overall times at 37° C and at the colder temperature were identical. By varying the temperature of the cold pulse it was possible to separate the change in the peak height and the delay as separate entities. 相似文献
6.
MASAHARU DOMON 《Development, growth & differentiation》1983,25(6):537-545
The cell cycles of the early cleavage stages of the mouse were analyzed by examining Feulgen-stained ova. The period from ovulation to the completion of second cleavage division was investigated. The ova donors were C57BL/6 × DBA/2 female mice, which were hormonally superovulated. To estimate the durations of DNA synthesis and mitotic phases of the cleavage divisions, the ova were pooled into culture medium, and as a function of time, aliquots were removed from the batch of pooled ova. The ova specimens were Feulgen-stained and classified as the ova nuclei in G1 , S, G2 or mitosis by use of a cytophotometric technique and then the durations of these phases were determined by probit analysis.
The pronuclear stage had a generation time of 11 hr, with a G1 phase of 6 hr and a short S phase of 1.7 hr. In contrast the two-cell stage had a generation time of 18 hr, with a G1 phase of 2 hr and an S phase of 3 hr. The duration of cleavage division also changed; the first cleavage division spanned 3 hr while the second spanned 1 hr. 相似文献
The pronuclear stage had a generation time of 11 hr, with a G
7.
Abstract. Multivariate analysis of the expression of cyclin proteins and DNA content has opened new possibilities for the study of the cell cycle. By virtue of their cell cycle phase specificity, the expression of cyclins may serve, in addition to DNA content, as another marker of a cell's position in the cycle, and provide information about the proliferative potential of cell populations. Several applications of the methodology based on bivariate analysis of DNA content v . expression of B, E and D type cyclins are reviewed: 1 expression of cyclins by individual cells during their progression through the cycle can be studied, using exponentially growing cells without the necessity of cell synchronization or other perturbations of the cycle; 2 cells having the same DNA content but residing in different phases of the cycle (e.g. G2 diploid v. G1 tetraploid) can be distinguished; 3 cell transition from G0 to G1 and progression through G1 (e.g. mitogen stimulated lymphocytes) can be assayed; 4 the population of proliferating cells can be distinguished from noncycling cells based on dual cell labelling with a G1 and G2 cyclin antibody; 5 cyclin restriction points can serve as additional cell cycle landmarks to map the point of action of antitumour drugs; 6 unscheduled expression of cyclins (e.g. the presence of cyclin B1 during G1 and S) can be detected in several tumour transformed cell lines, possibly indicating disregulation of the machmery of cell cycle progression. The last finding 6 is of special importance, because such disregulation may be of prognostic consequence in human tumours. 相似文献
8.
Abstract. Cellular uptake of [3 H]thymidine ([3 H]TdR) and incorporation into DNA of Ehrlich ascites tumour cells were studied in relation to the cell cycle by measuring the activity in the acid-soluble and insoluble parts of the cell material. Cells were synchronized at various stages of the cell cycle using centrifugal elutriation. The degree of synchrony of the various cell fractions was measured by flow-cytofluorometric DNA analysis. From the cellular uptake, the TdR triphosphate (dTTP) concentration of a mean cell in an unseparated cell population was calculated to be 20 × 10-18 mol/cell. The pool activity of G1 cells was unmeasurable but rose to maximum values at the border of the G1 -S phase. It decreased again during G2 . The [3 H]TdR incorporation into DNA was low during early S phase, reached a maximum value at two-thirds of the S phase and decreased again during late S phase. These changes in DNA synthesis were not due to changes in the dTTP pool being a limiting factor. During maximum DNA synthesis, 10%× min-1 of the dTTP pool was utilized, at which time the pool size also decreased by about 30%. Changes in pool size during the cell cycle have to be taken into account when the results of incorporation of radioactive TdR into DNA are discussed. 相似文献
9.
Abstract. The transit times of Chinese hamster ovary cells through the phases of their cell cycle were measured using dual parameter flow cytometry to measure DNA content and the presence of monoclonal antibodies to bromodeoxyuridine. Up to four separate populations can be accurately measured: unlabelled cells in G2 + M; labelled cells that have not yet divided; labelled cells that have already divided; and the unlabelled cells that were originally in G1 plus the cells that were originally in G2 + M and have since divided. The fractions of cells in these populations can be easily followed in time and the usual kinetic properties can be estimated from these fractions, or combinations thereof, including the times through G1 , S, G2 + M and the cycle time. We present equations for analysing this type of data and comment on which equations are most appropriate for measuring specific kinetic properties of the cells. 相似文献
10.
Abstract. Two methods involving labelling cells with bromodeoxyuridine (BrdUrd) have been used to study by flow cytometry the effect of hyperthermia (43°C for up to 1 h) on Chinese hamster V79 cells. One method involved the use of an antibody to BrdUrd after pulse-labelling the cells either before or at time intervals after treatment. In the second method, the cells were incubated continuously in BrdUrd after heat treatment, and the components of the cell cycle were then visualized by staining with a combination of a bis-bcnzimidazole and ethidium bromide. All three methods showed that heating at 43°C stopped DNA synthesis which, at 37°C, subsequently recovered reaching the normal rate 8–12 h later. The cells in S phase at the time of treatment then progressed to G2 where they were further delayed. Cells heated in G1 . after the recommencement of synthesis, progressed around the cycle, albeit slower than in unheated cells. The difference between the cells in G1 and S phases at the time of treatment may account for the greater sensitivity of S phase cells to hyperthermia. 相似文献
11.
Claire Huckins 《Cell proliferation》1971,4(2):139-154
The duration of the mitotic cycle and of its components was analysed for each of the six successive generations of differentiating spermatogonia (A1 , A2 , A3 , A4, intermediate and B), using radioautographed whole mounts of seminiferous tubules from testes of adult Sprague-Dawley rats. Cell cycles were determined from two successive waves of per cent labeled metaphases obtained during the period of 81 hr after a single dose of 3 H-thymidine. Except for the A1 spermatogonia, all spermatogonial types (A2 to B) had similar cell cycle durations of 41-42.5 hr and comparable pre-DNA synthesis phases (G1 ) of 11-13 hr. Although the combined duration of DNA synthesis (S) and the post-synthesis phase (G2 ) remained identical for all the cell types including A1 , there was a progressive lengthening of the S period at the expense of G2 during the process of spermatogonial maturation. This change was most marked during the transition from A1 to A3 spermatogonia when the S period increased from 14 hr to 21 hr, and the G2 phase shortened from 13 hr to 7.5 hr. This feature seems to be unique to germ cells and may be associated with an increasing amount of heterochromatin in the nucleus. Excluding the development of type A1 cells, the entire process of spermatogonial maturation lasted for 208 hr. Combined data on cell cycle times indicated that every 313 hr or 13 days, a new sequence of spermatogonial differentiation was initiated by the A1 cells. This was equivalent to the duration of one 'cycle' of the seminiferous epithelium as measured by other techniques. 相似文献
12.
13.
M. Ffrench † J. P. Magaud‡ M. Arzounian C. Souchier C. Charrin† P. A. Bryon 《Cell proliferation》1990,23(4):251-260
Abstract. In some cases of acute lymphoblastic leukaemia (ALL) the percentage of cells in G2 + M is higher than anticipated when compared with the percentage in S phase. This increase in G2 + M, as detected by flow cytometry measurement of DNA content, may be due to an accumulation of cells, either in G 2 or during the end of S phase; it may also be related to the existence of small tetraploid clones generally ignored by cytogeneticists. In order to identify possible subpopulations of cells with a DNA index ≥ 2-0, we have compared the results of a cytogenetic analysis to the G2 + M values. We have also studied the distribution of S phase cells in 24 cases of ALL by incorporating 5-bromodeoxyuridine, labelling the cells by indirect immunofluorescence, and analysing them by flow cytometry after propidium iodide staining. The distribution of cells during S phase was quantified: no accumulation of cells was ever observed at the end of S phase. The question of the existence of small tetraploid clones, G2 arrested cells or cells with a G2 elongation remains open. However, we feel that it is more probable that, in this pathology, an elongation of the duration of G2 occurs. 相似文献
14.
The G2 block is a major response of cells to DNA damage and seem to be induced independently of p53 status. It is thought that the G2 block has a protective function and allows cells to repair their DNA. The molecular events involved in the formation of the G2 block therefore are of great interest. We have used pentoxifylline, a potent G2 delay abrogator, to study the expression of an essential component of the mitosis promoting complex (MPF), cyclin B1. Cyclin B1/G2 ratios are used to show that irradiation induces a decrease in cyclin B1 expression and that pentoxifylline restores cyclin B1 expression to control level. This confirms that suppression of cyclin B1 plays a role in the formation of the G2 cell cycle delay, and that elevating cyclin B1 expression is part of the mechanism of action of pentoxifylline on G2 blocked cells. 相似文献
15.
D-type cyclins are involved in regulation of cell traverse through G1 primarily by activating the cyclin-dependent kinase 4 (CDK4) and targeting it to the retinoblastoma tumour suppressor protein. There is a vast body of evidence that defective expression of D-type cyclins is associated with tumour development and/or progression. Immunocytochemical detection of D cyclins combined with multiparameter flow cytometry makes it possible to measure the expression of these proteins in individual cells in relation to their cell cycle position without the need for cell synchronization. This approach was used in the present study to compare the cell cycle phase specific expression of cyclins D3 and D1 in human normal proliferating lymphocytes and fibroblasts, respectively, with nine tumour cell lines of different lineage. During exponential, unperturbed growth, expression of cyclin D1 in fibroblasts from donors of different age, or cyclin D3 in lymphocytes, was limited to mid-G1 cells: Less than 7% of the cells entering S phase or progressing through S and G2 were cyclin D positive. In contrast, expression of either cyclin D1 or cyclin D3 in tumour cell lines of different lineage was not limited to G1 phase. Namely, over 80% of the cells in S and G2 +M were cyclin D positive in eight of the nine cell lines studied. The data indicate that while expression of cyclin D1 or D3 in normal cells is discontinuous, occurring transiently in G1 , these proteins are expressed in some tumour lines persistently throughout the cell cycle. This suggests that the partner kinase CDK4 is perpetually active throughout the cell cycle in these tumour lines. 相似文献
16.
HANS-MARTIN SEYFERT 《The Journal of eukaryotic microbiology》1979,26(1):66-74
SYNOPSIS.
Under the growth conditions employed, the G1 macronucleus of Tetrahymena pyriformis HSM contains 7.4 × 10-12 g DNA, the G2 micronucleus 0.42 × 10-12 g. DNA content from the Tetrahymena thermophila macronucleus did not significantly differ from that of HSM, but the micronucleus contained about twice as much DNA as the micronucleus of the HSM cells. The T. thermophila macronucleus contained on average enough DNA for ˜ 35 haploid micronuclear copies. A new spreading technic allowed separation of macronuclear substructures from cells of late G2 to early G1 . Photometric determination of DNA content of 345 individual structures suggested the existence of 5 different-sized macronuclear structures with a DNA content corresponding to 2, 4, 8, and 16 × the basic values. Comparison of the DNA content of these structures with (a) mitotic micronuclear chromosomes and (b) meiotic micronuclear chromosomes of T. thermophila cells suggests that the 5 basic values of macronuclear structures derive from structures of micronuclear chromosomes. The micronuclear chromosomes of T. pyriformis may be oligotenic. It is suggested that these results further our understanding of macronuclear organization. 相似文献
Under the growth conditions employed, the G
17.
Elucidation of the DNA Synthetic Cycle of Entamoeba Spp. Using Flow Cytometry and Mathematical Modeling 总被引:2,自引:0,他引:2
JAMES A. DVORAK SEIKI KOBAYASHI DAVID W. ALLING CLAIRE W. HALLAHAN 《The Journal of eukaryotic microbiology》1995,42(5):610-616
ABSTRACT. We developed a method to study the DNA synthetic cycles of Entamoeba histolytica and Entamoeba invadens by flow cytometry (FCM) based on a preparative procedure to reduce both high levels of natural fluorescence and non-specific adsorption of fluorochromes. We modeled G1 , S, and G2 phases as a series of overlapping Gaussian curves. Both E. histolytica and E. invadens displayed G1 , S, and G2 proportions that are consistent with eukaryotic cell populations in exponential or stationary growth phase. Exponential phase E. histolytica populations contained a hypodiploid subset with a mass of about 20% less than the diploid value which we estimate by FCM to be 24 × 10-14 g DNA/cell. Exponential phase E. invadens populations contained a hypodiploid subset with a mass of about 6% less than the diploid value which we estimate by FCM to be 30 × 10-14 g DNA/cell. 相似文献
18.
Abstract— An analysis of the [3 H]DFP-labelled catalytic subunits of mammalian (bovine SCG) acetylcholinesterase (AChE, EC 3.1.1.7.) indicates a monomer molecular weight of 75,000. This is equivalent to the mass previously determined for the smallest active form and demonstrates that the globular, or G forms, are respectively monomeric (G1 form, 4S), dimeric (G2 form, 6.5S) and tetrameric (G4 form, 10S). In the tetrameric G4 form the catalytic chains are associated in dimers, by disulphide bonds.
The effect of reduction and proteolysis has shown that the dimeric form (G2 form, 6.5S) is readily reduced into G1 , while the tetramer G4 is very stable, being only dissociated by a combination of reduction and proteolysis by high concentration of trypsin. The asymmetric forms A12 (16S), A8 (13S) and A4 (9S) are not sensitive to reduction, but are readily dissociated by low concentrations of trypsin, into each other, progressively liberating isolated tetramers. We obtained essentially identical results with AChE preparations from rat brain or superior cervical ganglion. These observations support a general model for the quaternary structure of acetylcholinesterase molecular forms. 相似文献
The effect of reduction and proteolysis has shown that the dimeric form (G
19.
Abstract The effect of cortisol on the proliferation of the rainbow trout fibroblast cell line, RTG-2, was examined in synchronous and asynchronous cultures. When the transition from G1 to S was synchronized by restoring serum to serum-deprived cultures, the addition of cortisol at the time of serum restoration delayed the entry of cells into S phase. However, if cortisol was added 24 h after serum restoration, at the G1 /S transition point, the subsequent peak of DNA synthesis was unaffected. In asynchronous cultures cortisol inhibited [3 H]-thymidine and [3 H]-uridine but not [3 H]-leucine incorporation into acid-insoluble material. If the exogenous nucleoside concentration was raised, [3 H]-thymidine but not [3 H]-uridine incorporation continued to be inhibited by cortisol. This suggested that cortisol's effect on [3 H]-thymidine incorporation reflected a change in entry into S phase and not just on thymidine uptake and metabolism. Cortisol inhibited the proliferation of RTG-2 in asynchronous cultures. At 1000 ng/ml of cortisol a reduction in cell number became apparent before the RTG-2 cultures were confluent, whereas at 100 ng/ml the reduction only became evident in confluent cultures. The synthetic antiglucocorticoid, RU 486, which acts at the level of the corticosteroid receptor, blocked the growth inhibition by cortisol. These results suggest that cortisol regulates rainbow trout fibroblast proliferation via the corticosteroid receptor and that the G1 /S transition is one point at which this regulation occurs. 相似文献
20.
Sakumura T Fujii Z Umemoto S Murakami T Kawata Y Fujii K Minami M Sasaki K Matsuzaki M 《Cell proliferation》2000,33(1):19-28
The direct effects of the nucleoside transporter inhibitor dilazep on the cell cycle of mesangial cells have not before been investigated. The purpose of this study was to elucidate whether dilazep can inhibit the proliferation of mesangial cells and how it interferes with the cell cycle of these cells. DNA histograms were used and BrdUrd uptake rate was measured by flow cytometry. There was no significant difference in the cell numbers among the untreated group and the 10−5 M, 10−6 M or 10−7 M dilazep-treated groups at 24 h of incubation. However, at 48 and 72 h, the cell numbers in the dilazep-treated groups were significantly lower compared with that of the untreated group (P0.005). The DNA histograms of cultured rat mesangial cells at 12, 24, and 48 h of incubation with 10−5 M dilazep showed that the ratio of the S phase population in the dilazep-treated group decreased by 2.2% at 12 h, by 9.6% at 24 h, and by 18.9% at 48 h compared with the untreated group. The ratio of the G0 /G1 phase population in the dilazep-treated group significantly increased: 6.8% at 12h (P 0.05), 13.9% at 24 h (P 0.001), and 76.5% at 48 h (P 0.001) compared with the untreated group. A flow cytometric measurement of bivariate DNA/BrdUrd distribution demonstrated that the DNA synthesis rate in the S phase decreased after 6 h (P 0.005) and 12 h (P 0.05) of incubation compared with the untreated group. These results suggest that dilazep inhibits the proliferation of cultured rat mesangial cells by suppressing the G1 /S transition by prolonging G2 /M and through decreasing the DNA synthesis rate 相似文献