Studies on the vegetative life cycle of Chlamydomonas reinhardi Dangeard in synchronous culture III. Some notes on the process of zoospore liberation

When Chlamydomonas reinhardi cells liberate zoospores, theyexcrete into the medium a factor(s) which induces zoospore liberationof other cells that are not yet ready to liberate zoosporesby themselves. The "factor" is contained within cells at laterstages of the cell cycle, but its action is suppressed untilthe regular time of zoospore liberation in the cell cycle. (Received November 18, 1974; )     

Studies on the vegetative life cycle of Chlamydomonas reinhardi Dangeard in synchronous culture IV. Effects of 6-methyl purine on the revolution of the cell cycle

Cells of Chlamydomonas reinhardi Dangeard were synchronouslygrown under a 12 hr light-12 hr dark regime. The algal cellcycle under these conditions starts with a light-induced reaction(s)at the beginning of the light period and ends, after a definiteperiod of time (23–24 hr at 25°C), in zoospore liberation.When cells were exposed to 6-methyl purine for short periods(0.5–2.5 hr) at different times during the early and intermediatephases of the cell cycle, it exerted, as an analogue of adenine,two different effects on the revolution of the cell cycle: onea "lengthening" effect seen at its low concentrations in whichthe length of the cell cycle was somewhat prolonged, the othera "return to start" effect at higher concentrations. In thelatter a short exposure of cells to 6-methyl purine broughtthem to the starting point of the cell cycle concurrent withthe abortion of the cycle in process. When 6-methyl purine wasapplied during the later phase of about 1/4 the length of thecell cycle, it casued no effect. Control of the revolution ofthe algal cell cycle by an "adenine-involving reaction(s)" disturbedby this adenine analogue is discussed. (Received September 1, 1975; )     

Studies on the vegetative life cycle of Chlamydomonas reinhardi Dangeard in synchronous culture II. Effects of chloramphenicol and cycloheximide on the length of cell cycle

Cells of Chlamydomonas reinhardi Dangeard were synchronouslygrown under a 12 hr light— 12 hr dark regime. When thesecells were brought into contact with chloramphenicol for a shortperiod at early stages in the cell cycle, zoospore liberationwas delayed for a period which was nearly the same as that ofthe duration of contact with the antibiotic. When given at laterstages, the antibiotic caused no such effect. Cycloheximide,on the other hand, caused—when provided at some intermediatestage of the cell cycle— two different prolonging effectson the length of the cell cycle: one doubled the normal length(observed when the drug was administered at certain stages)and the other caused a delay similar to that caused by chloramphenicol.Interestingly, no prolonging effect was observed when cycloheximidewas given either at early stages or at later stages, such asduring the last 1/4 period of the cell cycle preceding zoosporeliberation. Based on these results, three phases were distinguishedin the algal cell cycle: "chloramphenicolsensitive", "cycloheximide-sensitive"and "insensitive" phases. Considering the known facts aboutthe modes of action of the two antibiotics inhibiting proteinsynthesis, discussions were made on the significance of proteinsynthesis in chloroplasts and in cytoplasm in determining thelength of the cell cycle. (Received October 12, 1970; )     

Cell Cycle Control in Arabidopsis

Although the basic mechanism of cell cycle control is conservedamong eukaryotes, its regulation differs in each type of organism.Plants have unique developmental features that distinguish themfrom other eukaryotes. These include the absence of cell migration,the formation of organs throughout the entire life-span fromspecialized regions called meristems, and the potency of non-dividingcells to re-enter the cell cycle. The study of plant cell cyclecontrol genes is expected to contribute to the understandingof these unique developmental phenomena. The principal regulatorsof the eukaryotic cell cycle, the cyclin-dependent kinases (CDKs)and cyclins, are conserved in plants. This review focuses oncell cycle regulation in the plant Arabidopsis thaliana . Whileexpression of one Arabidopsis CDK gene, Cdc2aAt, was positivelycorrelated with the competence of cells to divide, expressionof a mitotic-like cyclin, cyc1At, was almost exclusively confinedto dividing cells. The expression of the Arabidopsis -type cyclinsappears to be an early stage in the response of plant cellsto external and internal stimuli. Arabidopsis thaliana (L.) Heynh.; cell cycle; CDK; cyclin; plant development; plant hormone     

Studies on the cell wall of Chlorella I. Quantitative changes in cell wall polysaccharides during the cell cycle of Chlorella ellipsoidea

1. 1. The cell wall of Chlorella ellipsoidea was fractionated intotwo components, alkali-soluble hemicellulose and alkali-insoluble"rigid wall". The former was composed of several neutral sugars,i.e. rhamnose, xylose, arabinose, mannose and galactose, andthe latter had glucosamine as a main constituent sugar.
2. 2.Quantitative changes in both hemicellulose and "rigid wall"contents during the cell cycle were followed using synchronouslygrown cells. The two cell wall components showed markedly differentchanges. Hemicellulose increased in proportion to the enlargementof the cell surface area in the growing phase, while the "rigidwall" remained almost constant in this phase. The "rigid wall"increased only in the reproduction phase—the time of autosporeformation.

(Received September 26, 1977; )     

Returning Dark-Induced Cell Cycle to the Beginning of G1 Phase by Red Light Irradiation in Fern Adiantum Protonemata

In filamentous protonemata of Adiantum capillus-veneris L. preculturedunder continuous red light, the progression of cell cycle whichwas induced by transferring the protonemata to the dark wasinhibited and the phase returned to the beginning of G1 by redlight irradiation unless the cell cycle had progressed to a"point of no return" which coincides with the beginning of Sphase. The effect of red light was reversed by subsequent far-redlight. Typical red far-red reversibility indicates that thephotoreceptive pigment is phytochrome. (Received May 17, 1984; Accepted June 21, 1984)     

Exit from the Mitotic Cycle in Root Meristems of Zea mays L.

The choice between two modes of exit from the mitotic cycleat the margins of meristems has been made easier by surveyingthe range of the numbers of cell contacts between contiguousfiles in root apices of Zea mays L. The range shows that somecells must go out of cycle while others remain in cycle forat least three further generations. The view that cycling endsby a fall in the proliferative fraction is supported by theexistence of pulse-labelled telophases in the proximal regionof the menstem. These are most likely due to acceleration ofthe mitotic cycle which has to be contrasted with decelerationof the overall rate of cell proliferation. The work is discussedin relation to patterns of cycling in the different tissuesof the apex. mitotic cycle, cell size, meristem, proliferative fraction, Zea mays L, maize     

Studies on the vegetative life cycle of Chlamydomonas reinhardi Dangeard in synchronous culture I. Some characteristics of the cell cycle

Cells of Chlamydomonas reinhardi Dangeard were grown synchronouslyunder a 12 hr light-12 hr dark regime. Time courses of nucleardivision, chloroplast division, "apparent cytokinesis" and zoosporeliberation were followed during the vegetative cell cycle inthe synchronous culture. Liberation of zoospores occurred atabout 23–24 hr after the beginning of the light periodat 25°C. Four zoospores were produced per mother cell underthe conditions used. At lower temperatures, the process of zoosporeliberation as well as length of the cell cycle was markedlyprolonged, but the number of zoospores produced per mother cellwas approximately the same. At different light intensities,lengths of the cell cycle were virtually the same, while thenumber of zoospores liberated was larger at higher rather thanat lower light intensities. During the dark period, nuclear division, chloroplast divisionand apparent cytokinesis took place, in diis order, and proceededless synchronously than did the process of zoospore liberation.When the 12 hr dark period was replaced with a 12 hr light periodduring one cycle, the time of initiation as well as the durationof zoospore liberation was litde affected in most cases, whereasnuclear division, chloroplast division and apparent cytokinesiswere considerably accelerated by extended illumination. Whenalgal cells which had been exposed to light for 24 hr were furtherincubated in the light, zoospore liberation started much earlierand proceeded far less synchronously, compared with that under12 hr light-12 hr dark alternation. (Received October 12, 1970; )     

Cell cycle-dependent expression of volume-activated chloride currents in nasopharyngeal carcinoma cells

Patch-clamping and cell imageanalysis techniques were used to study the expression of thevolume-activated Cl current,I_Cl(vol), and regulatory volume decrease (RVD)capacity in the cell cycle in nasopharyngeal carcinoma cells (CNE-2Z). Hypotonic challenge caused CNE-2Z cells to swell and activated aCl current with a linear conductance, negligibletime-dependent inactivation, and a reversal potential close to theCl equilibrium potential. The sequence of anionpermeability was I > Br > Cl > gluconate. The Cl channelblockers tamoxifen, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB),and ATP inhibited I_Cl(vol). Synchronous cultures of cells were obtained by the mitotic shake-off technique and by adouble chemical-block (thymidine and hydroxyurea) technique. Theexpression of I_Cl(vol) was cell cycle dependent,being high in G₁ phase, downregulated in S phase, butincreasing again in M phase. Hypotonic solution activated RVD, whichwas cell cycle dependent and inhibited by the Cl channelblockers NPPB, tamoxifen, and ATP. The expression of I_Cl(vol) was closely correlated with the RVDcapacity in the cell cycle, suggesting a functional relationship.Inhibition of I_Cl(vol) by NPPB (100 µM)arrested cells in G₀/G₁. The data also suggest that expression of I_Cl(vol) and RVD capacity areactively modulated during the cell cycle. The volume-activatedCl current associated with RVD may therefore play animportant role during the cell cycle progress.

     

Cell cycle-related changes in transient K(+) current density in the GH3 pituitary cell line

Our aim was to determinewhether the expression of K⁺ currents is related to thecell cycle in the excitable GH3 pituitary cell line. K⁺currents were studied by electrophysiology, and bromodeoxyuridine (BrdU) labeling was used to compare their expression in cells thereafter identified as being in the S or non-S phase of the cellcycle. We show that the peak density of the transient outward K⁺ current (I_to) was 33% lower incells in S phase (BrdU+) than in cells in other phases of the cellcycle (BrdU). The voltage-dependence of I_towas not modified. However, of the two kinetic components ofI_to inactivation, the characteristics of thefast component differed significantly between BrdU+ and BrdU cells.Recovery from inactivation of I_to showedbiexponential and monoexponential function in BrdU and BrdU+ cells,respectively. This suggests that the molecular basis of this currentvaries during the cell cycle. We further demonstrated that4-aminopyridine, which blocks I_to, inhibited GH3cell proliferation without altering the membrane potential. These datasuggest that I_to may play a role in GH3 cellproliferation processes.

     

Proximal tubular epithelial cells are generated by division of differentiated cells in the healthy kidney

We searched for evidence for a contribution of stem cells in growth of the proximal S3 segments of healthy rats. According to the stem cell model, stem cells are undifferentiated and slow cycling; the bulk of cycling cells are transit amplifying, rapidly cycling cells. We show the following. 1) By continuous application of a thymidine analog (ThA) for 7 days, S3 proximal epithelial cells in healthy kidneys display a high-cycling rate. 2) Slow-cycling cells, identified by lack of ThA uptake during 14 days of continuous ThA application up to death and by expression of the cell cycle protein Ki67 at death, have the same degree of differentiation as quiescent cells. 3) To detect rapidly cycling cells, rats were killed at various time points after injection of a ThA. Double immunofluorescence for ThA and a cell cycle marker was performed, with colocalization indicating successive divisions. During one week after division, daughter cells display a very low proliferation rate, indicating the absence of rapidly cycling cells. 4) Labeling with cyclin D1 showed that this low proliferation rate is due to cycle arrest. 5) More than 50% of the S3 cells entered the cell cycle 36 h after a potent proliferative stimulus (lead acetate injection). We conclude that generation of new cells in the proximal tubule relies on division of differentiated, normally slow-cycling cells. These may rapidly enter the cycle under an adequate stimulus. immunohistochemistry; cell cycle; proliferation; renal stem cells; proximal tubule; renal epithelial cells     

Cell Cycle Changes in the Shoot Apex of Silene coeli-rosa During the Second and Third Days of Floral Induction

The cell cycle in Silene coeli-rosa shoot apices was measuredto test whether or not early components of the floral stimulus,produced during the 2nd and 3rd long days (LD) of an inductiveLD treatment, resulted in an increase in the duration of G₂phase in constant 20–24 h cell cycles. Plants were grownat 20°C in short days (SD) of 8 h light and 16 h darknessfor 28 d (day 0). Starting on day 0, plants were given SD or3 LD each comprising an identical 8 h day and 16 h photo-extension,or 3 dark-interrupted (d.i.) non-inductive LD, interrupted at1700 h of each day with 1 h of darkness. The cell cycle (percentagelabelled mitoses method) and changes in cell number were determinedin the shoot apical meristem. During days 1–2 of the SDtreatment, the cell cycle and mean cell generation time (MCGT)was 18 and 32 h, respectively, giving a growth fraction of 56%.During days 2–3, the cell cycle and MCGT shortened to15 and 23 h, respectively (growth fraction = 65%). During days1–2 of the LD and d.i. LD treatments, cell cycles andMCGTs were 9–10 and 27–29 h, respectively, resultingin smaller growth fractions (about 33%). Thus, shortened cellcycles and altered growth fractions occurred regardless of whetheror not the treatment was inductive. The LD treatment resultedin a marked shortening of G₁ and, to a lesser extent, S-phase,whilst G₂ remained constant. These changes were consistent withincreases in the proportion of cells in G₂ during the photoextensionof each LD which were suppressed during the comparable periodsof the d.i. LD treatment. The latter treatment resulted in eachphase occupying virtually identical proportions of the cellcycle as in the SD treatment. Thus, the unique cell cycle responsesto the initial part of the inductive LD treatment were increasesin the proportion of cells in G₂ coupled with G₁ and G₂ beingof similar duration. Cell cycle, mean cell generation time, shoot apex, Silene coeli-rosa     

Potassium channel expression level is dependent on the proliferation state in the GH3 pituitary cell line

Previously, we showed that the peakdensity of the transient outward K⁺ current(I_to) expressed in GH3 cells was different inthe S phase than in other phases of the cell cycle. Using cellsynchronization, we show here that I_to dropsprecisely at the quiescent (G₀ phase)/proliferating transition. This change is not due to a modification in the voltage dependence of I_to, but rather to a modificationin its inactivation kinetics. Molecular determination of K⁺channel subunits showed that I_to required theexpression of Kv1.4, Kv4.1, and Kv4.3. We found that the increase inI_to density during the quiescent state wasaccompanied by an increase in Kv1.4 protein expression, whereas Kv4.3expression remained unchanged. We further demonstrate that the linkbetween I_to expression and cell proliferation isnot mediated by variations in cell excitability. These results providenew evidence for the cell cycle dependence ofI_to expression, which could be relevant inunderstanding the mechanisms leading to pituitary adenomas.

     

Cell Cycle Timing of Replication of the Nuclear Gene Encoding Chloroplastic NADP-Specific Glutamate Dehydrogenase Isoenzymes in Chlorella sorokiniana

In synchronized Chlorella sorokiniana cells, the NH₄⁺ inducibleNADP-specific glutamate dehydrogenase enzyme (NADP-GDH) accumulatedin a linear manner throughout the first cell cycle. Early inthe following second cell cycle, an increase in its rate ofaccumulation occurred that was proportional to the increasein total cellular DNA in the previous cell cycle. In synchronizedbacterial cells, increases in rate of linear accumulation ofinducible enzymes coincide with the time of replication of theirstructural genes. To determine whether the rate change in NADPGDHaccumulation resulted from a delay in replication of its nuclearstructural gene (gdhN) in fully induced C. sorokiniana cells,the cell cycle timing of replication of this gene was comparedto that of another nuclear gene, nitrate reductase (nia), andof a chloroplast gene, ribulose bisphosphate carboxylase large-subunit(rbcL), in synchronized cells cultured in NH₄⁺ or NO₃^–(uninduced) medium. The gdhN and nia genes replicated withinthe period of nDNA synthesis and rbcL within the period of ctDNAsynthesis in cells growing in either nitrogen source. Therefore,the delayed rate change in enzyme accumulation results froma process that regulates expression of the gdhN gene after itsreplication. (Received July 16, 1994; Accepted November 28, 1994)     

家蚕细胞周期蛋白基因BmCcnl1的克隆及表达分析

家蚕Bombyx mori由受精卵到完成胚胎发育孵化的过程中, 细胞进行大量的分裂和分化, 然而滞育性卵的胚胎细胞分化至G2期便停滞在此阶段。为了探索这一发育阶段细胞内的分子调控, 本研究以人Homo sapiens的细胞周期蛋白基因cyclin L1为模板, 成功克隆了家蚕同源基因BmCcnl1（GenBank登录号: FJ889988）。BmCcnl1基因开放阅读框（open reading frame, ORF）全长1 254 bp, 编码417个氨基酸。利用Protean软件分析得出BmCcnl1蛋白预测分子量为49 kDa, 等电点为9.84。利用DNA重组技术构建了BmCcnl1基因的重组表达载体pET-21d-BmCcnl1, 对其进行原核表达, 其表达的蛋白以包涵体形式存在。利用RT-PCR技术分析了BmCcnl1基因在胚胎发育过程中的转录水平, BmCcnl1基因在非滞育性卵的胚胎发育阶段基本保持相对稳定的转录表达, 而滞育性卵从蛾体产下经过72 h后已经检测不到BmCcnl1基因的转录。结果提示, BmCcnl1基因与胚胎期滞育及非滞育性卵的发育调控相关。对该基因的克隆和表达分析为今后研究家蚕胚胎发育及细胞周期调控奠定了基础。     

Cell Cycle Activation During Imbibition and Visible Germination in Embryos and Megagametophytes of Jack Pine (Pinus banksiana Lamb.)

Flow cytometric determination of cell cycle activation duringimbibition and visible germination in five families of jackpine (Pinus banksiana Lamb.) embryos and megagametophytes revealedthat in seeds that had undergone no imbibition the majorityof cells were in the 2C state. As the imbibition period increased,less of the nuclei were blocked in the G0/G1 state and morebecome active in the cell cycle. The augmentation in the nucleiactive in the 2C–4C cycle as well as those with DNA levelshigher than the 4C state occured gradually and preceeded radicleemergence. In megagametophyte tissue examined at various stagesof imbibition, cell cycle activity became apparent rapidly followingimbibition. In nuclei of green and white embryos examined separatelythe ²frequency distributions were significantly different forall three families after 144h. As imbibition period increased,fewer nuclei from the green embryos were blocked in the 2C state,and more became active in the 2C–4C cell cycle. This wasnot the case for white embryos where no significant linear relationwas noted. Cell cycle activity in the hypocotyl+cotyledons regionand the emerging radicle were examined separately. Functionalrelations found in the hypocotyl+cotyledons region were notevident in the radicle. As visible germination proceeded, cellcycle activity in the hypocotyl + cotyledons region for thisperiod of germination showed a reversal of the activity notedduring imbibition: fewer nuclei were active and once again ahigher proportion were found in the 2C state. cell cycle; C levels; DNA content; flow cytometry; germination; imbibition; jack pine; megagametophyte; Pinus banksiana Lamb     

Identification of proteins interacting with the Arabidopsis Cdc2aAt protein

Cyclin-dependent kinases (CDK5) control the progression throughthe cell cycle. Using a two-hybrid approach, two clones encodingproteins interacting with the Arabidopsis thailana CDK Cdc2aAtwere identified. One clone encoded a novel putative substrateof Cdc2aAt, whereas the second clone was identified as a D-typecyclin (cycDl;1). Key words: Arabidopsis thaliana, cell cycle, cyclin, cyclindependent kinases, yeast two-hybrid screening     

SYNCHRONOUS CULTURE OF CHLORELLA: I. KINETIC ANALYSIS OF THE LIFE CYCLE OF CHLORELLA ELLIPSOIDEA AS AFFECTED BY CHANGES OF TEMPERATURE AND LIGHT INTENSITY

1. Using the technique of synchronous culture, investigationsweremade of the effects of temperature and light-intensityon cellularlife cycle of Chlorella ellipsoidea. Some improvementsin theculture technique for obtaining a good synchrony of algalgrowthwere described.
2. By following the changes of averagecell volume and cell numberoccurring during culturing, therates of the following processesof life cycle were determined:(i) "growth" (or the increasein cell mass) occurring from thestage of smaller cells (D_a)to the stage of ripened cell (L₃),(ii) "ripening" (or processofformation of "nuclear substances"as estimated from the averagenumber of daughter cells formedfrom single mother cell), and(iii) " maturing and division" which leads to the full maturationof mother cells (L-cells)and their division into separate daughtercells (D-cells).
3. "Growth"and "ripening" were found to be dependent in light,"maturingand division" light-independent. The time requiredfor "growth"and "ripening" (C) is dependent on temperaturebut independentof light intensity, the onset of "maturing anddivision" occurringat the same time (D) of culturing undervaried light intensities.The average cell volume at this stage(L₃),however, was foundto be markedly modified by light intensity;larger with highertemperatures (see Fig. 4).
4. Changes in incubation temperature(under the condition of saturatinglight intensities) were foundto affect the life cycle in thefollowing way: (i) The timeof onset of "maturing and division"(D), varies markedly withculturing temperature; earlier athigher temperatures, (ii)The average cell volume at this stagealso depends on temperature; smaller at higher temperatures.
5. The average number of daughtercells (n) emerging from singlemother cells, was found to beuninfluenced by culturing temperature;(4.0–4.1 underthe conditions of the present study). Itwas found that thedivision number n is remarkably varied bychanging the lightintensity in the "growth" and "ripening"phases; 2.0 at 1 kilolux,3.7 at 5 kilolux, 4.2 at saturatinglight intensities (10 and25 kilolux). This finding was explainedby assuming a light-dependentformation of "nuclear substances"during the "growth" and "ripening"phases, the quantity of thesubstances in the cell at L₃ stagedeterminig the division number.
6. The experimental data wereanalyzed reaction kinetically, therate constants and othercharacteristics of the reactions constitutingthe processesof life cycle were determined, and values forthe apparent activationenergy for each reaction were computed.The reactions were discussedwith special reference to theirrelationship with photosyntheticprocess was discussed.

(Received November 7, 1959; )     

Replicon Size, Mean Rate of DNA Replication and the Duration of the Cell Cycle and its Component Phases in Eight Monocotyledonous Species of Contrasting DNA C Values

Various aspects of the cell cycle were measured in the apicalmeristem of primary and seminal roots of eight monocotyledonousangiosperms: Oryza sativa (0.6 pg), Zea mays (2.4 pg), Pennisetumamericanum (2.5 pg), Aegilops umbellulata (5.1 pg), Hordeumvulgare (5.5 pg), Triticum monococcum (6.2 pg), Secale cereale(8.6 pg) and Tulipa kaufmanniana (22.6 pg), representing a 38-foldvariation in DNA C values. Using 4-d-old roots of the firstseven species and 21-d-old Tulipa roots, replicon size and ratesof replication were determined by DNA fibre autoradiography,and the duration of the cell cycle and its component phasesby the percentage labelled mitoses method. When tested withDNA C value, no significant relationships existed for repliconsize, rate of DNA replication or duration of G 1. Significantpositive linear relationships were found between DNA C valueand cell cycle duration, duration of mitosis and G2 durationwhen all data were tested, but not when the Tulipa data wereexcluded. The only characters significantly related to DNA C value whenthe Tulipa data were included or excluded were the durationof S-phase, and the ratio of the interval required for a repliconto replicate its allotted DNA (Rs) to the duration of S-phase(Ds). The Rs: Ds ratio is a measure of synchrony of repliconactivation, and the higher the DNA C value the lower this ratiobecame. We concluded that there was a nucleotypic effect ofDNA C value on this ratio and that the interval between activationof replicons became protracted and hence S-phase lengthenedas C value increased. Cell cycle, NA C value, DNA replication, replicon, S-phase