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1.
The process of continuous resynchronization with excess thymidine provides sufficient cell material for accurate chemical determination of DNA and RNA in HeLa S3 cells at hourly intervals during the cell cycle. Total DNA is constant during the non-S phase portion of the cell cycle but varies widely among cycles of synchronous growth. Total cellular RNA content increases linearly in the G1 phase and accelerates to a higher linear rate of accumulation, which remains constant during most of the S and G2 phases. The ratios of early and late cycle rates of RNA accumulation are not constant among cycles.  相似文献   

2.
Recent reports have shown that CDDP interacts with RNA and protein as well as DNA. We studied the alteration of cell cycle, cellular RNA content and the effect of nucleic acid metabolism on cultured cancer cells after treatment with CDDP by flow cytometry and 3H incorporation assay. The alteration of cell cycle was found to be accumulation of cells in after delay S phase in cytostatic concentrations, CDDP inhibited 3H-TdR uptake markedly at this time and 3H-UR uptake earlier. Increase in RNA content accompanied accumulation of cells in G2M phase. This increase was not a specific phenomenon caused by CDDP, because increase in RNA content was also induced by other inhibitors of DNA synthesis. It is more likely that the direct alteration of cell cycle and cellular RNA content due to action of DNA-combined CDDP rather than that of RNA-combined CDDP.  相似文献   

3.
The rates of uridine-5-3H incorporation into RNA and the rates of uridine uptake into the acid-soluble pool during the cell cycle of V79 Chinese hamster cells were examined. Cells cultured on Eagle''s minimal essential medium supplemented with fetal calf serum, lactalbumin hydrolysate, glutamine, and trypsin displayed rates of incorporation and uptake which increased only slightly during G1 and accelerated sharply as DNA synthesis commenced. In contrast, cells cultured on minimal essential medium supplemented only with calf serum exhibited rates of incorporation and uptake which increased linearly through both G1 and S. The transition from one pattern to the other can be induced within 24 hr and is completely reversible. The nonlinear pattern exhibited by cells grown on the supplemented fetal calf serum medium can also be overcome with high exogenous uridine concentrations. In the presence of 200 µM uridine, these cells display a linear pattern of increase in rates of uridine incorporation and uptake. It is concluded that at lower uridine concentrations the pattern of increase in the rate of uridine incorporation into RNA during the cell cycle for a given population of cells is dependent upon the rate of uridine entry into the cell, and that this pattern is not rigidly determined but can be modified by culture conditions.  相似文献   

4.
The rate of RNA synthesis in synchronously growing HeLa S3 cells was determined as a function of position in the cell generation cycle. Measurements throughout the cycle of both the rate of incorporation of radioactively-labeled uridine and of the total amount of RNA indicate that (1) the rate of RNA synthesis is constant (or increases only slightly) during G1, approximately doubles during the first half of S, and then remains constant during the remainder of S and G2, and (2) cells attain the average G1 rate of RNA synthesis very early in G1, and maintain the average G2 rate until mitosis. If the initiation of DNA synthesis is blocked, the acceleration of RNA synthesis is markedly reduced or eliminated. Further experiments in which DNA synthesis was inhibited at different times in S, or to varying degrees from the beginning of S, suggest that the extent to which RNA synthesis is accelerated depends on the amount of DNA duplicated. These data also indicate that duplication of the first half, and in particular the first few per cent, of the DNA complement results in a disproportionate acceleration of RNA synthesis. The possibility that fluctuations in the sizes of precursor pools may lead to misinterpretation of labeled-uridine incorporation data was examined. Experiments indicate that in this system pool fluctuations do not cause invalid measures of RNA synthesis. It is concluded that RNA synthesis occurs throughout interphase, but undergoes a two-fold increase in rate which is dependent on the duplication of DNA.  相似文献   

5.
During the annual three-phase growth cycle of white spruce [Picea glauca (Moench) Voss] the vegetative shoot apex changed in anatomical configuration. Relative amounts of DNA, histones, RNA, and proteins were measured in three cytohistological zones and were related to the anatomical changes during ontogeny. An extended period of DNA synthesis (S) and G2 preceded an increase in the number of apical initial cells which were part of the mammillary apex. While DNA and histones were generally synchronous during ontogeny, the ratio of DNA to histone increased on June 20. This loss of histone and subsequent increases in RNA and cytoplasmic proteins preceded the appearance of needle primordia on next year's apex. We propose that induction of the apex to reorganize and form needle primordia occurred when the DNA was in a 2C condition, following the loss of histone on June 20.  相似文献   

6.
The proliferating cells of mouse epidermis (basal cells) can be separated from the non-proliferating cells (differentiating cells) (Laerum, 1969) and brought into a mono-disperse suspension. This makes it possible to determine the cell cycle distributions (e.g. the relative number of cells in the G^ S and (G2+ M) phases of the cell cycle) of the basal cell population by means of micro-flow fluorometry. To study the regenerative cell proliferation in epidermis in more detail, changes in cell cycle distributions were observed by means of micro-flow fluorometry during the first 48 hr following adhesive tape stripping. 3H-TdR uptake (LI and grain count distribution) and mitotic rate (colcemid method) were also observed. An initial accumulation of G2 cells was observed 2 hr after stripping, followed by a subsequent decrease to less than half the control level. This was followed by an increase of cells entering mitosis from an initial depression to a first peak between 5 and 9 hr which could be satisfactorily explained by the changes in the G2 pool. After an initial depression of the S phase parameters, three peaks with intervals of about 12 hr followed. The cells in these peaks could be followed as cohorts through the G2 phase and mitosis, indicating a partial synchrony of cell cycle passage, with a shortening of the mean generation time of basal cells from 83-3 hr to about 12 hr. The oscillations of the proportion of cells in G2 phase indicated a rapid passage through this cell cycle phase. The S phase duration was within the normal range but showed a moderate decrease and the Gj phase duration was decreased to a minimum. In rapidly proliferating epidermis there was a good correlation between change in the number of labelled cells and cells with S phase DNA content. This shows that micro-flow fluorometry is a rapid method for the study of cell kinetics in a perturbed cell system in vivo.  相似文献   

7.
UV-induction of thymine dimers in cellular DNA and their excision during different phases of the cell cycle of HeLa S3 cells were studied. Induction of thymine dimers was higher in the mitotic phase and the middle of the S phase than in the G1 phase and from the late S phase to the early G2 phase which are rather insensitive to UV. However, there is no significant difference in excision rate of UV-induced thymine dimers from the irradiated cells through the cell cycle. These findings indicate that the cyclic variation of UV-survivals during the cell cycle may be due to differences in the amount of thymine dimers in cellular DNA induced by UV-irradiation.  相似文献   

8.
It has been reported that the response of target cells to steroid hormone (SH) stimulation may depend on their position in the cell cycle. The DNA and RNA contents of malignant cells of the endometrium cultured in vitro were measured using flow cytometry (FCM). We also measured estrogen receptor (ER) and progesterone receptor (PR) levels of cells at different positions in the cell cycle. The G1 and S phases of the cell cycle were investigated using cells synchronized by sodium n-butyrate (G1 block), methotrexate (S block), and excess thymidine (S block). For DNA measurements, the cells were stained with propidium iodide following RNase treatment. For RNA measurements (double-stranded RNA) the cells were treated with DNase. We found that S phase synchronization by methotrexate was 136.2% of control (100%). Using the excess thymidine block and release procedure, the S phase fraction was 185.1% of control. G1 phase synchronization by sodium n-butyrate was 134% of control. The estrogen receptor level in G1 phase synchronized cells increased to 5.94 fmol/micrograms DNA in the cytosol and 12.35 fmol/micrograms DNA in the nuclear fraction. These levels represent a sevenfold total increase over that of the control estrogen receptor level. Cells in S phase showed no significant increase in estrogen receptor levels over control cells. Based on this study, the functional increase of the steroid receptor was most significant in the G1 phase.  相似文献   

9.
The combined effect of mercury (HgCl2) and high temperature on the growth and synthesis of nucleic acid and protein, and on the cell cycle of HeLa S3 cells was investigated. The subsequent growth of the cells was dose-dependently inhibited by mercury at 37.2° and 41.2°C. The inhibitory effect of mercury on subsequent growth was enhanced at the higher temperature. IC50 values for DNA and RNA synthesis but not protein synthesis, at 41.2°C, were significantly lower than those at 37.2°C (P<0.05,P<0.01, respectively). Flow cytometric analysis using synchronous cells indicated the possibility of blocking of cell cycle progression in the early part of S phase by the combined treatment. These results suggest that the cytotoxicity of mercury to cell growth was enhanced at the higher temperature and that this enhancement is related to the increased inhibitory effect of mercury on DNA and RNA synthesis and on the cell cycle at high temperatures.  相似文献   

10.
SYNOPSIS. The size, composition and physiology of average cells have been studied in cultures of Acanthamoeba castellanii during the phases of logarithmic growth and population growth deceleration (PGD). Most of the features examined were relatively constant during log phase, but had significant changes during PGD. Average cell volume increased about 60% and total dry mass about 15–20% during the latter period. Total protein content remained constant thruout both growth phases, but cytochrome oxidase doubled during PGD. DNA, RNA and glycogen levels began to change during late log phase. DNA decreased about 50% and RNA increased about 75%. Glycogen decreased 50% during the RNA build-up and then increased to a plateau above the log phase level. A final decrease in glycogen followed an increase in the relative numbers of cysts in late PGD. It was found that PGD begins when O2 becomes limiting and evidence that the subsequent changes in macromolecule composition are related to encystation is discussed.  相似文献   

11.
12.
Summary Nuclease-colloidal gold complexes and silver staining were used to visualize intranucleolar nucleic acids and argyrophilic proteins of the nucleolar organizers in bud cotyledonary cells ofPisum sativum. In the G0–1 inhibited bud, a few RNA molecules were detected in the fibrillar component and in the unique fibrillar centre, close to the boundary with the fibrillar component of the nucleolus. DNA was present in the fibrillar component, in the fibrillar centre and in a few fibres crossing the perinucleolar halo. The acidic proteins were localized at the periphery of the fibrillar component but they were also present in the unique fibrillar centre. In the reactivated bud, RNA was particularly concentrated in the granular component and along fibres crossing the perinucleolar halo; a few RNA molecules were also detected at the boundary between the small fibrillar centres and the fibrillar component. DNA was localized in the same nucleolar component as in the inhibited bud, but it was distributed between several fibrillar centres. Acidic proteins coated these DNA loci. In the inhibited and reactivated bud connections between nucleolar DNA containing structures were displayed. The data are discussed in relation to the present knowledge of the functional architecture of the nucleolus.Abbreviations DNA deoxyribonucleic acid - DNase deoxyribonuclease - G0–1 phase G1 phase of the cell cycle indefinitely prolonged - PEG polyethylene glycol - RNA ribonucleic acid - RNase ribonuclease - S and G2 phases synthetic and postsynthetic phases of the cell cycle - SPB saline phosphate buffer  相似文献   

13.
We have measured the kinetics of specific globin mRNA and Friend virus (FV) RNA synthesis by hybridization to immobilized cDNA after induction of differentiation of two erythroleukemia cell lines (F4N, B8) by butyrate and Me2SO. The induction with butyrate in these cell lines occurs very rapidly (16–24 h). Cell cycle analysis was made of the populations throughout induction by flow cytofluorometry. The kinetics of commitment of cell populations to terminal differentiation by butyrate was determined by removal of inducer at various times and scoring of benzidine staining cells (hemoglobin producing). In addition, the cell cycle dependence of commitment was determined by flow sorting out of G1 and S+G2 cells various times after addition of inducer and scoring benzidine-stained colonies after growth in methylcellulose. Cells exposed to inducer were also sorted by cell cycle phase using an elutriator rotor. The amount of globin mRNA synthesis in the different cell populations was then determined.
1. 1. It was found that an 8–12 h period in butyrate was required before (a) globin specific mRNA was synthesized; and (b) commitment to differentiation occurred. The time course of globin mRNA synthesis was positively correlated with G1 arrest, as has been also found by others.
2. 2. The increase of FV RNA synthesis was not found during G1 arrest. It occurred early and before commitment.
3. 3. Commitment of cells to irreversible differentiation upon butyrate induction occurs only during the G1 phase of the cell cycle.
4. 4. Globin mRNA synthesis occurs first only in G1 cells.
5. 5. Globin mRNA is synthesized later in all phases of the cell cycle.
These data suggest that (a) commitment to differentiation and globin mRNA accumulation are coupled; and (b) that both events occur only in G 1 cells after a pre-commitment phase of about 12 h.  相似文献   

14.
The growth potential of turbot Scophthalmus maximus larvae and juveniles was studied using nucleic acid‐based indices and protein variables. The experiment was carried out from 4 to 60 days post hatching (dph). A significant increase in instantaneous growth rate during metamorphosis and retarded growth rate during post‐metamorphic phase were observed. Ontogenetic patterns of DNA, RNA and protein all showed developmental stage‐specific traits. The RNA:DNA ratio decreased up to 12 dph, then increased rapidly till 19 dph and fluctuated until 35 dph followed by a decline to the end. The RNA:DNA ratio was positively correlated with growth rate of juveniles during the post‐metamorphic phase, whereas this ratio was not a sensitive indicator of growth during the pre‐metamorphic phase and metamorphosis. The protein:DNA ratio showed a similar tendency to the RNA:DNA ratio. Changes of DNA content and protein:DNA ratio revealed that growth of S. maximus performed mainly by hyperplasia from 4 to 12 dph and hypertrophy until 21 dph during the pre‐metamorphic larval phase. Growth was dominantly hypertrophical from the early‐ to mid‐metamorphosing phase and hyperplastic thereafter. The results show that the DNA content and protein:DNA ratio can evaluate growth rates of larval and juvenile S. maximus on a cellular level.  相似文献   

15.
We examined replication of the autonomous parvovirus Aleutian mink disease parvovirus (ADV) in relation to cell cycle progression of permissive Crandell feline kidney (CRFK) cells. Flow cytometric analysis showed that ADV caused a composite, binary pattern of cell cycle arrest. ADV-induced cell cycle arrest occurred exclusively in cells containing de novo-synthesized viral nonstructural (NS) proteins. Production of ADV NS proteins, indicative of ADV replication, was triggered during S-phase traverse. The NS+ cells that were generated during later parts of S phase did not undergo cytokinesis and formed a distinct population, termed population A. Formation of population A was not prevented by VM-26, indicating that these cells were arrested in late S or G2 phase. Cells in population A continued to support high-level ADV DNA replication and production of infectious virus after the normal S phase had ceased. A second, postmitotic, NS+ population (termed population B) arose in G0/G1, downstream of population A. Population B cells were unable to traverse S phase but did exhibit low-level DNA synthesis. Since the nature of this DNA synthesis was not examined, we cannot at present differentiate between G1 and early S arrest in population B. Cells that became NS+ during S phase entered population A, whereas population B cells apparently remained NS- during S phase and expressed high NS levels postmitosis in G0/G1. This suggested that population B resulted from leakage of cells with subthreshold levels of ADV products through the late S/G2 block and, consequently, that the binary pattern of ADV-induced cell cycle arrest may be governed merely by viral replication levels within a single S phase. Flow cytometric analysis of propidium iodide fluorescence and bromodeoxyuridine uptake showed that population A cells sustained significantly higher levels of DNA replication than population B cells during the ADV-induced cell cycle arrest. Therefore, the type of ADV-induced cell cycle arrest was not trivial and could have implications for subsequent viral replication in the target cell.  相似文献   

16.
The methylation of nucleic acids has been investigated during the cell cycle of an asparagine dependent strain of transformed fibroblasts (BHK 21 HS 5). The synchrony was carried out by a partial asparagine starvation of cells for 24 hours. The amino acid supply induced all cells to enter synchronously the G1 phase. Methylation and DNA synthesis were respectively measured by pulsed [methyl-14C] methionine and [methyl-3H] thymidine incorporation. DNA methylation followed a biphasic pattern with maximal methyl incorporations during both S phase and mitosis. A partial desynchronisation induced the S phase of the second cycle to proceed before all the cells have achieved their division. Hydroxyurea was used in order to inhibit the DNA synthesis of cells entering the second cell cycle, which might interfer with the mitosis of the first one. The inhibitor was added either at the first beginning of cell division or during all the G1 phase. In both conditions it suppressed 3H thymidine incorporation of the second cycle. However, mitosis took place and methylations occurred as in previous experiments. The DNA methylation of the mitotic phase in the first cell cycle could thus be dissociated from the classical post-synthetic DNA maturation and did not correspond to any DNA methylation appearing in the course of the second cell cycle.  相似文献   

17.
18.
The cell membrane potential of cultured Chinese hamster cells is known to increase at the start of the S phase. The putative role of the cell membrane potential as a regulator of cell proliferation was examined by following the cell cycle traverse of synchronized Chinese hamster cells in the presence or absense of high exogenous levels of potassium. An increase in external potassium levels results in a depressed membrane potential and a reduced rate of cell proliferation. A potassium concentration of 115 mM was used in experiments with synchronized cells since at that level cell proliferation is almost completely halted, recovery of growth is rapid and complete, and the membrane potential is reduced to a level well below that normally found in cells in the G1 phase. A mitotic population was divided into four aliquots and plated in either control medium or medium containing 115 mM K+. Cells placed directly into high K+ medium were retarded in their exit from mitosis and displayed a delayed and abnormal entry into the S phase. If control medium was added after two hours, cell cycle traverse was normal, but delayed by two hours compared to control cells. If the mitotic cells were plated directly into control medium and two hours later were shifted to high K+ medium, the cells entered the S phase in the absence of the normally observed increase in membrane potential and proceeded to the next mitosis normally. It was concluded that the increase in membrane potential observed at the start of the S phase in isolated synchronized cells is not a requirement for the initiation of DNA synthesis. In addition, sensitivity to the high potassium regimen was found at two different times during the cell cycle. In one case, cells were impeded in their transit through mitosis. Such cells displayed an altered chromosome structure which may account for the partial mitotic block. In the second case, synchronized cells displayed a sensitivity to the high potassium regimen in early G1 which appeared to be separate from the block in mitosis and independent of a change in the membrane potential.  相似文献   

19.
Cell heterogeneity during the cell cycle   总被引:7,自引:0,他引:7  
  相似文献   

20.
Functional and optimal activities of the (Na+-K+)ATPase, as determined by ouabain-sensitive K+ influx in intact cells and ATP hydrolysis in cell homogenates respectively, have been measured during the cell cycle of neuroblastoma (clone Neuro-2A) cells. The cells were synchronized by selective detachment of mitotic cells. The ouabain-sensitive K+ influx decreased more than fourfold from 1.62 +/- 0.11 nmoles/min/10(6) cells to 0.36 +/- 0.25 nmoles/min/10(6) cells on passing from mitosis to early G1 phase. On entry into S phase a transient sixfold increase to 2.07 +/- 0.30 nmoles/min/10(6) cells was observed, followed by a rapid decline, after which the active K+ influx rose again steadily from 1.03 +/- 0.25 nmoles/min/10(6) cells in early S phase to 2.10 +/- 0.92 nmoles/min/10(6) cells just prior to the next mitosis. The ouabain-insensitive component rose linearly through the cycle in the same manner as the protein content/cell. Combining total K+ influx values with efflux data obtained previously showed that net loss of K+ occurred with transition from mitosis to G1 phase while net accumulation occurred with entry into S. Throughout mid-S phase net K+ flux was virtually zero, but a large net influx occurred again just before the next mitosis. The (Na+-K+)ATPase activity measured in cell homogenates decreased rapidly from mitosis to G1 phase and increased steadily throughout S phase, but the transient activation on entry into S phase was not observed. Complete inhibition of the (Na+-K+)ATPase mediated K+ influx by ouabain (5 mM) prevents the cells from entering S phase, while partial inhibition by lower concentrations of ouabain (0.2 and 0.5 mM; km = 0.17 mM) causes partial blockage in G1 and, to a lesser extent, a reduced rate of progression through the rest of the cell cycle. We conclude that the transient increase in (Na+-K+)ATPase mediated K+ influx at the G1/S transition is a prerequisite for entry into S phase, while maintenance of adequate levels of K+ influx is necessary for normal rate of progression through the rest of the cell cycle.  相似文献   

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