Nuclear DNA Metabolism in the Tip of the Cortical Strands of Viscum album L

The tips of the cortical strands of Viscum album were investigatedby autoradiographic and cytophotometric methods. It is shown that DNA synthesis and mitotic activity of the sub-apicalmeristematic cells are constant during the whole year. Theirnuclear DNA content varies from 2C to 4C values. The elongated cells which cover the meristematic zone are characterizedby no DNA synthesis, no mitotic activity and a 2C nuclear DNAcontent. They are ‘blocked’ in the presyntheticphasc G₁ of their cellular cycle, without polyploidy. The relationship between polyploidy and secretion is discussed. Viscum album, mistletoe, nucleus, DNA, polyploidy     

Commitment of Mitotic Cells to Meiosis during the G2 Phase of Premeiosis

In the developing anther, archesporial cells that proliferateby mitotic division are converted into meiotic cells duringthe premeiotic interphase. Experiments with explanted microsporocytesof Lilium and Trillium were made to obtain evidence for theconversion of mitotic to meiotic cells during the premeioticperiod. Explanted premeiotic cells were cultured through thedivision cycle at relatively high division frequencies and showeda variety of division types with respect to chromosomal events.The type of division depended on the premeiotic stage at whichthe cells were explanted. Cells in the G₁, S and early G² phasesunderwent mitotic division and formed a diad or binucleate monad.Cells explanted at the late G₂ phase were cultured throughoutthe normal meiotic cycle, which resulted in typical tetrad configuration. In microsporocytes explanted during the main part of the G₂interval, centromere behavior was meiotic, but chromosome pairingand chiasma formation were disturbed. Thus, she G₂ intervalwas shown to be critical for the commitment of mitotic cellsto meiotic division. Detailed analysis showed that the intracellularchanges that commit the cells to meiosis begin shortly aftercompletion of premeiotic DNA synthesis and that these changesare progressive and cumulative. (Received February 2, 1982; Accepted May 24, 1982)     

The Quiescent Centre and Rates of Mitosis in the Root Meristem of Allium sativum

The root apices of Allium sativum have been examined by continuous-and pulse-labelling with tritiated thymidine and by colchicinetreatment to measure the time parameters of the mitotic cyclein various parts of the meristem. There is a quiescent centre of 30–50 cells whose averagerate of mitosis is low because the G₁ period is extended toabout 140 h compared with about 4 h in the othe regions of themeristem. The stele just above the quiescent centre and at 200microns above it and the cap initials just below the quiescentcentre are very similar in their mitotic cycles, the total lengthsof which are about 30 h of which nearly half is taken up byDNA synthesis. Allium thus differs from Zea in having root capinitials whose mitotic cycle is not telescoped by the eliminationof the G₁ phase. These facts are discussed in relation to theradiosensitivity of the meristem.     

Synthesis of DNA during Mitosis

Growing roots of Zea mays in tritiated thymidine for brief periodsconfirms that G₁ may be eliminated from the mitotic cycle andDNA synthesis may be advanced into telophase but no further,in the fastest dividing cells of the cap initials, but not inthe stele.     

Mitotic Activities and Levels of Nuclear DNA in the Apical Meristem of Silene armeria (strain SI.2) Following Application of Gibberellin A3

Strain S1.2 of Silene armeria was grown under an 8h-photoperiodand treated with GA₃ every day for 20 days. This growth substancecaused stem elongation, but no flowering in this long-day plant.Changes in the mitotic index and DNA content of cells in thevarious zones of the apical meristem before, during and afterGA₃ treatment were described. Mitotic activity and increasein the proportion of nuclei at the 4C level (S+G₂ phase of thecell cycle) were strongly stimulated in the rib-meristem, andto a lesser extent in the lateral zone, but not in the axialzone. This stimulation of apical activity reached a peak aftertwo GA₃ treatments and then declined gradually, so that after20 days the activity in GA₃-treated meristems was lower thanthat in untreated controls; at this point most cells were inthe G₁ phase. When the GA₃ treatment was discontinued, there was a gradualincrease in the mitotic activity which ultimately reached thesame level as that in controls. Stem elongation ceased and leavesformed aerial rosettes. It is concluded that in vegetative plants of strain S1.2 ofSilene armeria GA₃ acts mainly on the rib-meristem cells whichresults in stem elongation. Lack of response in the axial cellsexplains why GA₃ fails to induce flowering in this strain ofSilene armeria. (Received June 18, 1983; Accepted August 3, 1983)     

Flow Cytometric Determination of Nuclear Replication Stages in Tomato Seeds during Priming and Germination

Flow cytometric determination of DNA levels in embryos of fullymatured dry tomato (Lycopersicon esculenium) seeds revealedlarge amounts of 2C DNA signals, indicating that most cellshad arrested in the cell cycle at the presynthetic G₁ phaseof nuclear division. After imbibition in water, an augmentationof the 4C signal in the embryonic root tip region was found.This increase could be ascribed to cells entering the syntheticphase of nuclear division leading towards the doubling of chromosomalmaterial. In the root tip cells, 4C:2C ratios increased I dafter imbibition in water though radicle emergence started 2d later. Apparently, DNA synthesis preceded germination. Onlya small increase in the number of cells with 4C DNA levels wasfound in the rest of the embryonic tissues. In whole dry seeds,DNA histograms revealed both a 2C signal and a considerable6C peak, the latter originating from the endoreduplicated endosperm. A priming period of 14 d in PEG-6000 considerably enhanced therate and uniformity of germination. In the ungerminated seeds,the 4C DNA signal of root tip cells started to increase after3 d incubation in PEG. The ratio of 4C:2C steadily increasedduring the 14 d priming period, though did not reach the levelobtained after hydration in water. Upon priming, the 4C:2C ratiowas constant after redrying the seeds towards the original moisturecontent, indicating that the chromosomal material in the rootcells had stably ceased cell cycle activity at the G² phase.The present results indicate that the beneficial effects ofpriming on seedling performance are associated with the actionof replicative DNA synthetic processes prior to germination. Lycopersicon esculeniumMill, tomato, DNA content, flow cytometry, priming, seed, nuclear replication stage, C levels     

Amounts of Nuclear DNA and Internal Morphology of Gibberellin- and Abscisic Acid-deficient Tomato (Lycopersicon esculentum Mill.) Seeds During Maturation, Imbibition and Germination

Using X-ray photography and flow cytometry, the internal morphologyand DNA replication activity of wild type (wt), GA- (gib-1 )and ABA-deficient (sit^w ) tomato (Lycopersicon esculentum Mill.cv. Moneymaker) mutant seeds were studied. During seed formation,from 30 to 45 d after pollination (DAP) the endosperm becomessolid and the seed starts to gain desiccation tolerance. Atthis time significant changes occur in the amounts of DNA inradicle tip cells. At 30 DAP, radicle tip cells of the threegenotypes manifest about 60% of 2C, 30% of 4C and 10% of 8Camounts of DNA. Upon maturation (45 DAP onwards), most cellsin the seeds of the three genotypes arrest in the G₁phase ofthe cell-cycle with 2C amounts of DNA. However, a relativelyhigh proportion of cells with 4C amounts of DNA was detectedin the radicle tip cells ofsit^w compared with wild type andgib-1. At the well-matured stage (60 DAP), there were about 2% ofseeds with free space in wild type andgib-1 , and about 13%insit^w . At the over-matured stage (75 DAP), even more seedswith free space were found insit^w , whereas no increase in theproportion of the seeds with free space was detected in theother two genotypes. In -1.0 MPa PEG-6000 with or without 10µM GA₄₊₇, no germination occurred in well-matured wildtype andgib-1 seeds, whether or not they were dried after harvest.However,sit^w seeds were able to germinate both in over-maturefruit and in -1.0 MPa PEG-6000. Priming of dried seeds in -1.0MPa PEG induced a large amount of free space in almost all seedsof the three genotypes, and nuclear DNA synthesis in the radicletip cells of wild type andsit^w seeds. However, PEG priming offresh (non-dried) seeds had no effect on the amount of freespace and 2C/4C DNA ratios in wild type orgib-1 seeds, but didinduce free space in about 20–25% ofsit^w seeds and provoked4C signals insit^w seeds. Removal of the endosperm and testaopposite the radicle tip of seeds resulted in root protrusion,the induction of free space and an increase of 4C DNA signalsin the three genotypes. It is concluded that ABA is crucialfor the efficient arrest of tomato embryo radicle tip cellsin G₁phase upon maturation, whereas GAs play an important rolein re-initiating 4C DNA levels upon germination. Dormancy; flow cytometry; free space; Lycopersicon esculentum ; maturation; priming; seed; tomato     

Priming and accelerated ageing affect L-isoaspartyl methyltransferase activity in tomato (Lycopersicon esculentum Mill.) seed

Damage and degradation of cellular proteins is observed duringage-induced seed deterioration. L-Isoaspartyl protein methyltransferase(EC 2.1.1.77 [EC] ) is an enzyme hypothesized to play a role in limitingand repairing age-induced damage to proteins. Tomato (Lycopersiconesculentum Mill. ‘New Yorker’) seeds were assayedfor changes in L-isoaspartyl methyl-transferase activity duringaccelerated ageing and after osmotic priming. Accelerated ageingof seeds for 1–4 d at 45C and 100% relative humidityreduced germination from 94% to 71%, increased the mean timeof germination (MTG) from 2.4 to 5.8 d, and was accompaniedby a correlative decrease in L-isoaspartyl methyltransferaseactivity (r²=0.90). Aged and untreated seeds were primed for7 d at 20C in darkness using aerated solutions of 3% KNO₃ orpolyethylene glycol 8000 (PEG) with equivalent osmotic potential(–1.25 MPa). Priming with KNO₃ decreased the MTG, butdid not improve germination percentage for untreated seeds.Priming did not affect L-isoaspartyl methyltransferase activityin untreated seeds, but restored activity in aged seeds primedin KNO₃ to levels near that of untreated seeds. Priming withPEG did not effectively improve the MTG or increase L-isoaspartylmethyltransferase activity. During germination, L-isoaspartylmethyltransferase activity remained constant for 48 h post-imbibitionand then declined, suggesting that the enzyme was developmentallyregulated and inactivated or degraded as radicle emergence occurred. Key words: L-Isoaspartyl methyltransferase, protein repair, seed priming, accelerated ageing, Lycopersicon esculentum     

Effect of nickel at high concentration on proliferation of quiescent center cells and initiation of lateral root primordia in wheat seedlings

DNA synthesis and cell divisions in the quiescent center as well as initiation of lateral root primordia were investigated in the course of incubation of the roots of 3-day-old wheat (Triticum aestivum L.) seedlings on the medium with 0.1 mM NiSO₄ for 72 h. It was found that the earliest effect of nickel on proliferation of the quiescent center cells was associated with an increase in the mitotic index 6 h after the beginning of its action. This effect was assumed to depend on an increase in mitosis time. Twelve hours after the beginning of the effect of nickel, mitotic index became somewhat lower, and in 18 h it sharply decreased. Some dividing cells were observed among the initial cells of certain tissues and near the quiescent center even in 72 h. The portion of DNA synthesizing cell sharply decreased in 12 h, and in 48 h such cells were lacking. The main mechanism governing the termination of cell proliferation in the quiescent center as well as in the meristem and calyptrogen of the cap is the inhibition of cell transition to DNA synthesis. The cells that had time to start DNA synthesis or already finished it and were in other phases of the cycle continued a slow progression through the cycle and completed it. Sister cells, produced as a result of divisions, left the mitotic cycle in the phase G₁ and transited to dormancy. Nickel did not inhibit initiation and development of lateral root primordia. Resumption of DNA synthesis and cell divisions occurred not only in the pericycle and endodermis participating in the initiation of lateral root primordia but also in the cortex cells in the vicinity of developing primordia. In 18 h after the beginning of the experiment when the rate of the root growth considerably decreased, the region, where primordia were initiated, was located closer to the root tip. Subsequently, when elongation of the cells was inhibited, this region moved closer to the tip until structural disturbances occurred in the nuclei of the endodermal cells located near the root tip and elongated under the effect of nickel. The results concerning the effect of nickel and other heavy metals on root cell proliferation obtained by other researchers and the role of pericycle organization in the translocation and accumulation of nickel in the tissues are discussed.     

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)     

The Nuclear Endoreduplication Cycle in Metaxylem Cells of Primary Roots of Zea mays L.

The nuclear DNA content of metaxylem cells in roots of Zea mayscv. Golden Bantam reaches 16C or 32C by successive rounds ofDNA endoreduplication. Each phase of endoreduplication (endo-S)is separated by a non-DNA synthetic phase (endo-G). These phasesseem to occur in zones at fixed distances from the root tip.The duration of the phases in two of the endoreduplication cycles(4C–8C, 8C–16C) has been estimated in two ways.The first makes use of the rate of movement of cells throughthe positions along the root where the different phases of thecycle are occurring, the second uses labelling with methyl-[³H]thymidineand autoradiography. Both methods indicate that the endo-S phaseswhich cause the nuclear DNA content to rise from 4C to 8C andfrom 8C to 16C last 8–10 h, and that the intervening endo-Gphase lasts 8–12 h. DNA endoreduplication keeps pace withthe increase of nuclear volume; cell volume increases at a morerapid rate, however. Comparison of the endoreduplication cyclein the metaxylem with the mitotic cycle in the adjoining filesof parenchyma cells shows that the mitotic cells complete theircycle more slowly. DNA synthesis, endoreduplication cycle, mitotic cycle, root apex, Zea mays     

Physiological Studies on Germination of Phaseolus mungo Seeds: II. GLUCOSE AND ORGANIC-ACID METABOLISMS IN THE EARLY PHASES OF GERMINATION

The metabolism of glucose and organic acids was studied in theearly phases of germination of Phaseolus mungo seeds. Duringthe first 5–6 h of imbibition, ethanol fermentation activelyoccurred and in the following stage the TCA cycle seemed tobe markedly activated. CO₂ fixation seemed to be low, at leastin the early stage of germination. Aspartic acid was convertedactively into malic acid. This seemed to be a cause of the activesynthesis of malic acid observed.     

Influence of Environmental Stress on the Germination of Anabaena vaginicola Akinetes

Germination of akinetes of Anabeana vaginicola v. fertilissimaPrasad in response to environmental stress was studied. Additionof nitrate to the medium induced early and maximum germination(96 per cent), whereas less than half of the akinetes germinatedwhen either nitrate or phosphate was omitted from the medium.The pH range over which germination occurred was 7.0–9.0.The desiccated akinetes after rehydration germinated after acertain lag period, depending upon the dehydration state. Thetemperature optimum for germination and vegetative growth wasthe same (25 °C) and germination did not occur at 5 °Cor above 35 °C. The limit of heat shock tolerated was 55°C for 4 min. In addition to white light, only the red partof the visible spectrum induced germination. Ultraviolet radiationreduced germination rate presumably by inducing thymine dimersin DNA. The photoreactivating system (s) in akinetes is certainlynon-photosynthetic. LD₅₀ photon flux densities were 300 Jm^–2for akinetes and 240 Jm^–2 for vegetative cells. Anabaena vaginicola, blue-green alga, akinete, germination, environmental stress     

Inhibition of growth and photosynthesis of freshwater phytoplankton by ultraviolet A (UVA) radiation and subsequent recovery from stress

The effect of ultraviolet A (UVA) on growth and photosyntheticrate was studied in diatoms (Melosira spp.) of the phytoplanktonof a eutrophic lake and a cultured green alga Chloretla ellipsoidea.The cells were incubated under photosynthetically active radiation(PAR) (–UVA) or PAR + UVA conditions (+UVA). Growth ofC.ellipsoidea was retarded under +UVA, as shown by an increasein the lag period, but the rate of exponential growth was almostthe same in + and –UVA conditions. The photosyntheticrate was depressed markedly by UVA in Chlorella cells grownunder –UVA. In contrast, cells grown in +UVA showed onlyslight inhibition by UVA and after exposure to UVA for 6 daysthere was no inhibition. During the growth experiment, the cellularchlorophyll a content was higher in +UVA than +UVA grown cells.A similar effect was observed in diatoms from the eutrophicLake Suwa. In vivo fluorescence with (F_a) and without 3-(3,4-dichloropheny)-l,l-dimethylurea (DCMU) (F_b) and the photosynthetic rate were measured forC.ellipsoidea and the diatoms for 5 h under + and –UVAconditions. Soon after C.ellipsoidea had been subjected to +UVA,F_b and F_a / F_b decreased quickly and reached minima after 40min and 1 h, respectively. The suppressed in vivo fluorescenceresumed and full recovery was achieved after 4 h. This suggeststhat reactivation of the photosystem is acquired under prolongedexposure to UVA. A similar shift of F_a + F_b, but no change inFb, was found in diatoms by exposure to UVA. Changes in photosyntheticoxygen evolution by C.ellipsoidea under +UVA were similar tochanges in F_a + F_b. Degradation of chlorophyll a extracted inmethanol was enhanced by UVA. The rate of degradation by UVAwas independent of temperature from 15 to 34°C, suggestinga photochemical reaction. The results indicate that C.ellipsoideaand Melosira spp. acclimatize to prolonged UVA exposure by reactivationof the photosystem and enhanced cellular chlorophyll a synthesis.The ecological importance of these results to phytoplanktonproductivity in natural aquatic environments is discussed.     

The in vitro response of thymic lymphocytes of the pig to phytohemagglutinin

The response of thymic lymphocytes of the pig to phytohemagglutinin was studied with H³ thymidine in cultures, from 0–72 hours. At the beginning of the culture period 6–18% of lymphocytes were in DNA synthesis. during the first 24 hours a sharp decrease in the number of DNA synthesizing cells was observed in both pha and control cultures, although pha cultures consistently showed small but significantly greater numbers of DNA synthesizing cells. this was followed by a definite peak in DNA synthesis and mitotic response of a minority of the cells in pha cultures between 48–54 hours, whereas in control cultures activity ceased. in addition, a small proportion of the progeny of initially DNA synthesizing medium sized lymphocytes was apparently stimulated by pha and found in mitosis by 48 hours. It was concluded that the thymus contains a fraction of lymphocytes, not in the mitotic cycle, which are capable of being transformed by pha to mitotic activity. the data also suggests some stimulation of cells already in the mitotic cycle.     

The Effects of Cell Cycle Parameters on Cell Wall Regeneration and Cell Division of Cotton Protoplasts (Gossypium hirsutum L.)

Protoplasts of cotton cotyledons were isolated and culturedto undergo cell wall regeneration and cell division. DNA contentand cell cycle parameters of nuclei from cotyledons and/or protoplastswere determined by flow cytometry. The DNA content of cotton,Gossypium hirsutum L., was estimated to be 4·34±0·12pg DNA per nucleus. There was a strong positive correlation between G₂ or Sand G₂,and cell wall regeneration and cell division and a strong negativecorrelation between G₁, and cell wall regeneration and celldivision of cotton cotyledon protoplasts. The cell cycle statusof cotyledons changes during their development; as the cotyledonsenlarge, the proportion of cells in G₀ and G₁ phases of thecell cycle increases. The implication of these results in relationto protoplast growth and development is discussed. Key words: Cell cycle parameters, cell wall regeneration, cell division, flow cytometry, Gossypium     

THE ONSET OF MITOSIS AND DNA SYNTHESIS IN ROOTS OF GERMINATING BEANS

The time of onset of mitosis and DNA synthesis has been determined in roots of germinating seeds of Vicia faba. Mitosis is not initiated in all roots simultaneously. Dividing cells are seen 36 hr from the beginning of germination, but they are present in low frequency (0.02%). Dividing cells do not become frequent, i.e., occurring as 5% or more of all cells, until 56 hr, and it is not until 66–68 hr that all roots in a sample of 10 are mitotically active. DNA synthesis shows a similar sporadic beginning. It occurs in a few cells by 28 hr, and by 40 hr all roots exposed to ³H–thymidine show active incorporation. For most cells in these germinating roots DNA synthesis precedes mitosis. In one root in 10, however, some cells are unlabeled when they enter mitosis, indicating that they had spent the dormant period in the G₂ phase of the mitotic cycle. The presence of these cells determines whether or not roots show chromatid and chromosome aberrations following irradiation during germination.