A. González-Fernández

Duration of the division cycle in binucleate and mononucleate cells

Abstract The material used consisted of meristematic cells from the root-tips of onions ( Allium cepa ) grown in aerated tap-water at a temperature of 25 °. By using caffeine to label the cells (1-hr treatment with a solution of 0.1 per cent), a population of binucleate cells was induced, through inhibition of cytokinesis in the cells which were undergoing division at the time of the treatment. The development of the labelled population showed that the binucleate cells have a division cycle of 14 hr and the mononucleate cells a cycle of 13.5 hr. In meristematic cells, interphase lasts 11.2 hr and mitosis 2.3 hr, divided into: prophase, 64 min; metaphase, 18 min; anaphase, 13 min; and telophase 42 min.

Effect of caffeine and adenosine on G2 repair: mitotic delay and chromosome damage.

Proliferating plant cells treated during the late S period with 5-aminouracil (AU), give the typical response that DNA-damaging agents induce, characterized by: an important mitotic delay, and a potentiation of the chromosome damage by caffeine post-treatment. The study of labelled prophases, after a tritiated thymidine pulse, allowed evaluation of the mitotic delay induced by AU as well as its reversion by caffeine, while chromosome damage was estimated by the percentage of anaphases and telophases showing chromosomal aberrations. Post-treatment with adenosine alone has shown no effect on mitotic delay or chromosomal damage. However, when cells after AU were incubated in caffeine plus adenosine, the chromosome damage potentiation was abolished without affecting the caffeine action on mitotic delay. As a consequence, we postulate that caffeine could have two effects on G2 cells with damaged DNA: the first, to cancel their mitotic delay and the second to inhibit some DNA-repair pathway(s). Only this last effect could be reversed by adenosine.

Effect produced by inhibitors of RNA synthesis on mitosis.

Abstract The study of mitosis in root-tip cells of Allium cepa and in populations of synchronous binucleate cells induced in this material by treatment with caffeine enabled the authors to demonstrate the detention of cells in prophase caused by treatment with 3′ deoxyadenosine (3′AdR), ethidium bromide or ribonuclease. Cells incubated at the late interphase in presence of the drugs mentioned reached the prophase but, when observed over a period of 10 h, did not succeed in initiating the metaphase. Similarly, those cells which were in prophase at the beginning of the treatments remained in prophase. When treatment was applied to cells at metaphase, mitosis followed its normal course, and the subsequent mitotic stages were not prolonged. The results obtained suggest that a synthesis of RNA, probably specific, required to enable the cells to go on from the prophase to the metaphase of mitosis, takes place during prophase.

Duration of the division cycle in diploid, binucleate and tetraploid cells

Abstract By means of a caffeine treatment (0.1 per cent solution, 12–15 hr, at 25 °C) a binucleate cell population is induced in root-tips of Allium cepa. In the division of these cells (bimitosis) blocks of three cells, diploid-binucleatediploid (2n; 2n + 2n; 2n), are formed and nuclear fusion in the binucleate cells give rise to blocks diploid-tetraploid-diploid (2n; 4n; 2n), which development has been studied. Knowing the duration of the diploid cycle (13.5 hr), the tetraploid cell cycle was calculated about 14.5 hr. In short, the duration of the average cycle should be: for the mononucleate diploid cells, 13.5 hr; for the binucleate cells, 14 hr; and for the tetraploid cells, 14.5 hr. Therefore, the cycle time does not only depend on the DNA amount per cell, but also on the DNA distribution in one or two nuclei and perhaps the modifications of the ratio nuclear-surface area to nuclear-volume might explain the results obtained.

A model for cell cycle and growth kinetics in roots

A model is proposed which accounts for the cell cycle and growth kinetics in roots growing under steady state conditions. In the model the root is conceived as a single file of cells and the three sequential zones of the root are considered. Nc cells, arranged in a single longitudinal file, make up the meristem zone. The most distal cell of this file is the initial and it has an indefinite cycle capacity, whereas all the others are derived cells with a limited programme of n cycles. The model predicts Nc = 2n. The number (Nc) of meristem cells will allow us to calculate the number of cycles that any derived cell undergoes before arresting division and triggering off its elongation. The cycle kinetics are linear and therefore the distribution of the Nc cells in the cycle compartments is directly proportional to each phase duration, and constant with time. The rate of meristem cell formation is defined by the cell flow (ϕ) as the frequency of cells that pass any point of the cycle per time unit. It also represents the frequency of cells that the meristem supplies to the elongation zone per time unit. The elongation zone is formed by Ntr cells in the process of enlargement. The model predicts that N tr = ϕ . N o . T tr Ttr being the time that cells devote to their enlargement throughout this zone. The equation allows us to estimate this value. Changes in root length will depend only on length changes in the mature zone where G = N o . ϕ . L G being the rate of growth and L the final cell size. During symplastic growth the root may be regarded as a bundle of files with identical growth rates. This leads to quantitative relationships between the growth components of the different kinds of files. Lastly, the fitness of the single file model against the actual growing root is discussed.

Protein synthesis requirements at specific points in the interphase of meristematic cells

Abstract A 6 h treatment with anisomycin at a concentration of 1 μg/ml enables us to modify the steady-state kinetics of a meristematic cell population of Allium cepa , and this points to a difference of sensitivity to inhibition of protein synthesis between the several periods of the cell division cycle (G1, S, G2, M). The results show that the cells are incapable of entering the S period in the presence of the inhibitor, but that, where DNA synthesis has already been initiated, the synthesis continues in the cells in question. It was found, moreover, that there is a point in the early G2 period, which has a duration of approx. 3 % of the total duration of the cycle, at which the synthesis of specific proteins appears to determine the progression of cells to mitosis.

Mechanism of mitotic synchronization induced by 5-aminouracil

Abstract The mechanism of mitotic synchronization induced by 5-aminouracil (5-AU) in onion root meristems has been investigated. We studied the effect of the chemical upon asynchronous meristematic cells, as well as upon a synchronous fraction of them the position of which in the cell cycle was known. The kinetic studies have shown a fairly linear relationship between the treatment time and the induced mitotic wave. Therefore, cell accumulation in a particular zone of interphase appears to be the cause of mitotic synchronization. This zone seems to be located at the end of S period. 5-AU induces maximal mitotic delay on the cells located in the accumulation zone and severely depresses the S-G2 transit while the passage through G 2 remains relatively unaffected. The possibility that preferential inhibition affects the late replicating DNA synthesis is discussed.

Interphase development and beginning of mitosis in the different nuclei of polynucleate homokaryotic cells

The degree of synchrony in the course of the interphase periods G1, S and G2 and in the initiation of mitosis in the several nuclei of each cell of a polynucleate population induced by treatment with 0.1% caffeine, in root meristems of Allium cepa, through inhibition of cytokinesis in two successive cell divisions is analysed by means of labelling with 3H-thymidine.—The S period is initiated simultaneously in all the nuclei of each polynucleate cell, which supports the hypothesis of a factor present in the cytoplasm that is responsible for inducing DNA synthesis.—However, all the nuclei in a polynucleate cell do not pass from the S period to the G2 period simultaneously, those surrounded by the greatest amount of cytoplasm, generally the outer nuclei, being the first to complete the S period (“early nuclei”) and beginning the prophase before their fellow-nuclei in the same cell (“late nuclei”).—From the metaphase onwards, however, all the nuclei in a polynucleate cell continue to develop synchronously. The synchronizing mechanism has a twofold aspect: the shortening of the G2 period in the “late nuclei” and the lengthening of it in the “early ones” and, on the other hand, an arrest of prophase in the “early nuclei” until the “late ones” have caught up, which suggests the existence of an inhibiting factor produced by the “late nuclei” capable of acting upon the early ones through the cytoplasm.

A model for dynamics of cell division cycle in onion roots

SummaryThe study of the cell division cycle by means of caffeine labelling inAllium roots, at 15° C, employing intact root and decapitated roots at several levels (0.5, 1.0, 1.5, 2.0, and 2.5 mm) has shown that the number of cycles developed by the cells is constant at each meristem level. This number and the durations of the cycles are not affected by the decapitation. It is suggested that the cell cycle is controlled in the meristematic cells by an intracellular programme which would be developed throughout the meristem.However, the larger the region decapitated is, the more decreases the growth rate of the roots. The removal of the root cap (about 0.5 mm) did not modify the rate of root growth, although it blocked the geotropic response. The quiescent center is proposed as a source of auxin controlling cell elongation.

Cytokinesis at prophase in plant cells treated with ethidium bromide

Abstract The process of mitosis was arrested at commencement in root-tip cells of Allium cepa by treatment with ethidium bromide, although cytokinesis was apparently not affected and the cell plate was formed in the case of cells whose nuclear development was blocked in the prophase. The results suggest that cytokinesis is not dependent upon the synthesis of nucleic acids when once the cell has initiated the prophase.