CHANGES IN GLYCERALDEHYDE 3-PHOSPHATE DEHYDROGENASE ACTIVITY IN SHOOT APICAL MERISTEMS OF BRASSICA CAMPESTRIS DURING TRANSITION TO FLOWERING

An assay system has been developed for the histochemical determination of glyceraldehyde 3-phosphate dehydrogenase (G3PD) activity to indicate glycolytic pathway capacity during evocation in the shoot apical meristem of Brassica campestris L. G3PD activity was differentially distributed in a zonate pattern within the meristems at the vegetative, the transition, and the floral stages. The activity of G3PD changed in all apical zones of evoked apices, but especially in the central and the peripheral zones of apices at the prefloral stage. In the prefloral stage of development heavy enzyme activity was localized in island-like areas within the peripheral zone. These results indicate that 1) the capacity of glycolysis fluctuates during evocation, 2) during floral evocation the capacity of the glycolytic pathway parallels the capacity of the citric acid cycle and the electron transport system only at the prefloral stage, and 3) G3PD activity marks incipient floral primordia. It is proposed that the enzymic marking of an incipient floral primordium indicates the end of evocation in Brassica.     

A QUANTITATIVE STUDY OF CELLULAR EVENTS IN THE SHOOT APICAL MERISTEM OF BRASSICA CAMPESTRIS (CRUCIFERAE) DURING TRANSITION FROM VEGETATIVE TO REPRODUCTIVE CONDITION

Vegetative plants were induced to flower by 16-hr-long days. Apical buds were collected at intervals during several developmental phases up to 63 hr. A stereologic analysis and mitotic index study was conducted on median longitudinal sections of shoot apical meristems. A rise in the mitotic index occurred between 12 and 24 hr within central, peripheral and pithrib meristem zones. Preceding the floral stage a second increase in the mitotic index was observed in peripheral and central zones, but not in the pith-rib meristem zone. A significant rise in apical volume, cell number, height, and width began in the transitional stage and continued to the floral stage. Significant correlation coefficients were observed between these apical parameters. Relative volume and cell population of each zone remained constant from the vegetative to the reproductive stage. Volume fraction occupied by the nucleus and nucleolus remained constant within each zone during the same time period. In each zone the volume of the nucleus was significantly correlated to volume of the nucleolus. It appears a pre-inflorescence apex, while larger, is structurally similar to a vegetative apex.     

CHANGES IN MITOTIC ACTIVITY AND CELL SIZE IN THE APICAL MERISTEM OF HELIANTHUS ANNUUS L. DURING THE TRANSITION TO FLOWERING

Cellular changes in the shoot apical meristem of Helianthus annuus L. have been investigated in relation to its progress towards flowering. During the strictly vegetative phase, lasting for 6–7 days from sowing, mitotic divisions were confined to the peripheral zone, while the central mother cells zone proper, together with the distal cells positioned above the mother zone and also the central portion of the tunica, were relatively quiescent. Mitotic activity increased in the distal cells zone on day 8 and reached the level of that in the peripheral zone by day 12. This was accompanied by an enlargement of this zone and the consequent recession of the mother zone away from the central tunica. At the same time there was a substantial increase in the amount of cytoplasm in the cells of the central tunica. Mitotic activity in the central tunica began on day 12 and reached a peak on day 16. This zone then lost its distinct entity and was replaced by a uniform dome-shaped meristematic layer that became apparent by day 16. The cells of the mother zone remained quiescent during the transition period.     

A HISTOCHEMICAL STUDY OF THE SEEDLING SHOOT APICAL MERISTEM OF PINUS LAMBERTIANA

Vernalized seeds of Pinus lambertiana were scarified and planted in perlite. At 5, 8, 10, 13 and 16 days after planting, seedlings were selected for morphological examination and histochemical study. The shoot apical meristem consisted of a relatively homogeneous population of cells at 5 days. Cytohistological zonation was observed in the meristem by the eighth day and needle primordia initiation began at this time. Acid phosphatase (AP) activity was high in the extreme tip of the apex at 5 days. At 8 days AP activity was intense in the peripheral zone but weak in the apical initial and central mother cell zones. The apical meristem of the 10–16-day-old seedlings exhibited high AP activity in the peripheral zone only during the early stages of needle primordia initiation. The distribution of cytoplasmic and nuclear protein-bound SH was correlated with cytohistological zonation. Protein-bound SH was distributed relatively uniformly at 5 days, but by the eighth day the 4 cytohistological zones contained differential quantities. Succinic dehydrogenase (SD) activity was observed throughout the apex at 5 days, but by the eighth day the apical initial and central mother cell zones exhibited differentially greater levels of SD activity. Irradiation with 500 R of X-rays at 7 days after planting completely inhibited needle primordia initiation and disrupted the cytohistological zonation of the apex. Correlated with the inhibition of needle primordia initiation was the loss of SD activity in the apical initial and central mother cell zones. Irradiation also resulted in the gradual loss of protein-bound SH from the cytoplasm of the apical initial, central mother cell and peripheral zone.     

VARIATION IN CELL-CYCLE TIME AND NUCLEAR DNA CONTENT IN THE APICAL MERISTEM OF HELIANTHUS ANNUUS L. DURING THE TRANSITION TO FLOWERING

The mitotic cycle in the apical meristem of Helianthus annuus L. has been investigated during the transition to flowering. Towards the end of the strictly vegetative phase 8 days after sowing the average cell-cycle time, measured by colchicine-induced metaphase accumulation, was 37 hr in the peripheral zone, 83 hr in the central zone and 118 hr in the rib meristem. By Day 12 the cycle had shortened in all zones. By the time of floral initiation on Day 16 the cycle time had returned to its original value in the peripheral zone and the rib meristem, while in the central zone it continued to shorten to 33 hr, approaching the cycle time of the peripheral zone. Cytophotometric measurements of nuclear DNA showed that mitotic activation of the central zone was not associated with any reduction in the proportion of nuclei with a 4 C DNA content. It was calculated that the spatial and temporal variation in cell-cycle time was mainly a function of the length of the G₁/G₀ phase which lasted about 19 hr in the peripheral zone, 82 hr in the rib meristem, and declined from 55 to 21 hr in the central zone.     

STEREOLOGICAL STUDY OF ULTRASTRUCTURAL CHANGES IN THE SHOOT APICAL MERISTEM OF NICOTIANA TABACUM DURING FLORAL INDUCTION

Shoot apices of a short-day sensitive line of Nicotiana tabacum L. cv. NCTG-22 have been examined by electron microscopy for ultrastructural changes which occurred in the central zone over a 17-day period during the transition from vegetative to reproductive growth. Plants were grown in controlled-environment chambers of the NCSU Phytotron and exposed to an inductive photoperiod after a 6-wk juvenile phase. Ultrastructural changes were investigated from photomicrographs using a semi-automatic stereological procedure and a microcomputer. After exposure to only one inductive cycle cell and nuclear cross sectional areas in short-day plants were significantly larger than in long-day controls. Subsequently, under short days cross sectional areas of cells, nuclei, vacuoles and proplastids decreased, while mitochondrial cross sectional area and relative volume increased during the course of the induction period. In induced apices as cross sectional areas and relative volumes of vacuoles and proplastids decreased, their profile numbers increased. The reduction in cross sectional areas of cells and most organelles was associated with an increase in rate of leaf initiation and size of the apical dome. The demand for sufficient energy input to maintain the surge in growth and activity preceding floral initiation was reflected by the significant increases in cross sectional area, profile numbers and density of the mitochondria population. Even though the transition period is quite long for Nicotiana, cells and organelles in the central zone were observed to progress through similar changes in morphology that are known to occur in Sinapis and Xanthium which exhibit a more rapid and absolute response to photo-induction.     

芸薹属植物发育过程中顶端结构的解剖学研究

本文采用解剖学方法研究花椰菜、青花菜、结球甘蓝和大白菜在生长发育过程中顶端分生组织结构的变化及之间存在的差异。结果显示它们的顶端分生组织结构都是由最初幼苗的原套-原体结构逐渐发育到过渡型分区结构、典型化五个分区结构,至开始进入生殖生长时期的四个分区结构(形成层状细胞区消失)。四种植物在进入生殖生长后,顶端分生组织细胞行为不同:大白菜和甘蓝顶端亚外套两侧细胞分裂分化形成顶生叶原基,在顶生叶原基内侧的细胞将进行分裂产生花序侧枝原基。花椰菜和青花菜顶端亚外套两侧细胞分裂形成花序分生组织,花序分生组织增生即为花球体;内部解剖结构表现为分生组织不断分裂增多的过程。这些结果为研究花序表型发生的解剖学本质及分子生物学研究分生组织发育方向奠定了基础。     

CELL DIVISION STUDIES OF THE SHOOT APEX OF DATURA STRAMONIUM DURING TRANSITION TO FLOWERING

Seedlings of Datura stramonium L., although not photoperiodically sensitive, are useful for floral transition studies when raised in a growth chamber at a constant temperature of 25 C with a photoperiod of 8 hr of light (1,600-2,000 ft-c) and 16 hr of darkness. A terminal flower is formed after the seventh or eighth leaf primordium is produced. A constant rate of leaf initiation up to the time of flowering enables specific apical stages to be obtained and studied. Changes in the mitotic index, substantiated with calculated rates of cell division (measured by the accumulation of metaphases following treatment with colchicine) were studied in shoot apical zones during transition to flowering. Fluctuations in the mitotic index of each zone in the vegetative and transition apex with respect to apical stage as well as time of day were not statistically significant. The mitotic index of the summit zone of the vegetative apex was significantly lower than in the other zones whose mitotic indices were not significantly different from one another. During floral transition the mitotic index of the summit zone as well as the central zone (just below the summit zone) significantly increased while no significant changes were detected in the flank zones. It was shown that the mitotic index could be considered representative of the rates of cell division in Datura.     

CELL DIVISION KINETICS IN THE SHOOT APICAL MERISTEM OF CERATOPTERIS THALICTROIDES BRONG. WITH SPECIAL REFERENCE TO THE APICAL CELL

The duration of mitosis and the cell cycle were determined for defined cell populations of the shoot apical meristem of Ceratopteris thalictroides Brong. by using the colchicine-induced metaphase accumulation technique. The results indicate that the apical cell is mitotically active and cycles at an apparently greater frequency than the cells of subjacent populations. Duration of mitosis was similar for all cells of the meristem. These results are correlated with mitotic indices of control apices, the geometry of the apex, and the mean number of cells in the meristem. Shoot apices from adult plants were examined to determine mitotic indices within the meristem; mitotic activity was again noted for the apical cell. These results contradict recent proposals that the pteridophyte apical cell serves as a unicellular quiescent center which lacks histogenic potential and offer experimental support for the classical concept of apical cell function in those fern shoot meristems which terminate in a single apical cell.     

IN VITRO DEVELOPMENT OF THE ISOLATED SHOOT APICAL MERISTEM OF ANGIOSPERMS

Development of complete plants was achieved from isolated shoot apical meristems of Nicotiana tabacum L., Daucus carota L., Nicotiana glauca Grah., Tropaeolum majus L., and Coleus blumei Benth. The explants consisted of only meristematic dome tissue with no visible leaf primordia. A simple nutrient medium composed of the Murashige and Skoog salt mixture, 100 mg/liter myo-inositol, 0.4 mg/liter thiamin-HCl, 1-2 mg/liter IAA, 30 g/liter sucrose, and 1% agar was adequate. Histologically there occurred principally tissue enlargement during the first 3-6 days, followed by appearance of bipolar organization in 6-9 days and formation of a well-defined root apex and initiation of first leaf primordium by 12 days.     

MITOTIC ACTIVITY IN THE ROOT APICAL MERISTEM OF AZOLLA FILICULOIDES LAM., WITH SPECIAL REFERENCE TO THE APICAL CELL

The meristematic activity of the apical cell and its derivatives (merophytes) in the unbranched, determinate roots of Azolla filiculoides Lam. was investigated. The plane of division of the apical cell indicates that it is the initial of each merophyte. The division plane of each newly formed merophyte is strictly periclinal to the root surface and provides confirmation that the immediate derivatives of the apical cell cannot be the ultimate root initials. The frequency of cell division as determined by the mitotic index, and by the duration of the cell cycle as determined by the colchicine method, confirmed the meristematic activity of the apical cell. As roots increase in length, the duration of the cell cycle in the total meristem increases, with the apical cell possessing the longest cell cycle, whereas the immediate derivatives maintain approximately the same cycle duration as in shorter roots. In determinate Azolla roots, cell division appears to play a major role up to a certain root length, then increase in length is produced mainly by cell elongation.     

ON THE RELATIONSHIP OF LIVER GLUCOSE-6-PHOSPHATASE TO THE PROLIFERATION OF ENDOPLASMIC RETICULUM IN PHENOBARBITAL INDUCTION

The differentiated effects of phenobarbital treatment on liver microsomal enzymes have been further studied. The relationship between the resulting decrease in the specific glucose-6-phosphatase activity and the enhancement of formation of endoplasmic reticulum membranes with high drug-hydroxylating activity has been investigated with biochemical and histochemical methods. Biochemically and histochemically demonstrable glucose-6-phosphatase activity was found to be present in all endoplasmic reticulum membranes, including the phenobarbital-induced smooth-surfaced proliferates, even though there was an over-all decrease in activity. Actinomycin D did not inhibit the decrease in glucose-6-phosphatase activity. The findings are discussed with reference to the enzyme-membrane relationship in phenobarbital induction.     

香榧营养苗端细胞组织分区的超微结构观察

本文研究了榧树(Torreya grandis)成熟植株在季节生长中营养苗端的超微结构变化。各区域细胞的主要区别特征为:顶端原始细胞与亚顶端细胞相接的细胞壁较厚,液泡多分布于细胞游离面,质体中淀粉粒较小;亚顶端细胞壁较厚,液泡较大,质体中淀粉粒较大而多;周缘区细胞质体多不具淀粉粒,液泡也较小,胞间连丝丰富;肋状区细胞被大量的含淀粉质体及液泡占据了大部分空间,胞间连丝丰富。在季节变化的四个时期中,各区域细胞的亚显微结构特征亦不相同。休眠期各区细胞淀粉质体较发达,细胞壁较厚,液泡大;叶扩展期淀粉质体减少或消失;芽鳞形成期出现大量小液泡;新的顶芽形成期液泡增加,核糖体含量较高。讨论了各区域细胞核形态与其细胞活跃性的关系。     

HORMONAL REQUIREMENTS OF EXCISED DIANTHUS CARYOPHYLLUS L. SHOOT APICAL MERISTEM IN VITRO

Whereas a medium containing kinetin alone enabled a few Dianthus caryophyllus L. apical meristem dome explants to develop into rooted plants, the highest frequency of plants was obtained in one containing supplements of both IAA and kinetin. In an unsupplemented medium, continued development required that explants have 2 pairs of primordial and a pair of expanding leaves. Kinetin alone caused production of many new leaves, but the development was significantly less than when it was furnished in combination with IAA. IAA given alone caused meristem explants to develop primarily callus, roots, and a few leaves. Gibberellin and abscisic acid were without promotive effects on leaf and shoot formation. A balance of hormonal substances, synthesized in young leaf structures and relocated to the meristem, is proposed as the fundamental mechanism that regulates new leaf initiation in the shoot apex.     

AN ELECTRON MICROSCOPIC EVALUATION OF CYTOHISTOLOGICAL ZONATION IN THE SHOOT APICAL MERISTEM OF PINUS BANKSIANA

Shoot apical meristems of jack pine (Pinus banksiana) were examined by light and electron microscopy. Cytohistological zonation was evident when meristems were fixed in Craf IV, embedded in paraffin, and stained with Chlorazol Black E. When meristems were fixed for electron microscopy the cytoplasm of the apical initials and central mother cells each contained numerous lipid bodies and their nuclei contained little, if any, heterochromatin. The cytoplasm of the peripheral zone was rich in ribosomes. The nuclei of the peripheral zone and rib meristem were heterochromatic. Thus, the lack of heterochromatin in the nuclei and the dissolution of lipids in the cytoplasm of the apical initials and central mother cells appeared to contribute most to the organization and appearance (cytohistological zonation) of the shoot apex when standard histological techniques are used.     

EXPERIMENTAL STUDIES OF THE SHOOT APICAL MERISTEM OF SEED PLANTS. I. MORPHOLOGICAL AND CYTOCHEMICAL EFFECTS OF IAA APPLIED TO THE EXPOSED MERISTEM OF LUPINUS ALBUS

Application of 1.5 μg indoleacetic acid (IAA) in a lanolin droplet to the exposed apical meristem of Lupinus albus seedlings caused: (1) axillary buds to form closer to the apex than normal, (2) displacement of primordia formed during the first two plastochrons following treatment, and (3) significant increases in concentration of RNA, protein, and unsaturated lipids in the meristems. Primordial displacement tended to be random relative to the site of the treatment, which may be a feature common to dicotyledonous plants exhibiting spiral phyllotaxis. That IAA conferred initiation site capabilities to all of the peripheral zone for a short time was indicated by (2) and (3) above, and by decreases in concentrations of the observed compounds toward control levels after the second plastochron following treatment. Effects of IAA on RNA suggest that nucleic acid metabolism, and possibly gene action, was involved in the response. Kinetin or gibberellic acid had no apparent morphogenetic effect on Lupinus meristems.     

THE PICEA ABIES SHOOT APICAL MERISTEM IN CULTURE. I. AGAR AND AUTOCLAVING EFFECTS

Shoot apical meristems of Picea abies seedlings can be cultured on a relatively simple, defined, basal medium. Dome-like explants initially about 200 μ tall, without externally obvious primordia, and having dry weights of about 3 μg, usually initiate 5–10 new primordia within a week. They typically show 10- to 30-fold dry weight increases in three weeks. None of the 5,000 meristems cultured has produced any basal callus. Growth is strongly influenced by both the type and concentration of agar used to gel the medium. Dry weight yield increases as agar concentration decreases. This is probably partly due to increased diffusion rates of enzymes or other large molecules through more dilute agar gels but possibly also partly ascribable to unknown agarborne inhibitors. About half of the agar concentration effect can be eliminated by substituting glucose and fructose for sucrose in the medium. This suggests that diffusion of invertase through the agar gel in this medium may be a growth limiting factor. Growth of cultures is also promoted by autoclaving sucrose in the presence of the agar. The basis of this effect is not yet understood.     

A MORPHOMETRY STUDY OF THE ULTRASTRUCTURE OF ECHINOCEREUS ENGELMANNII (CACTACEAE). II. THE MATURE,ZONATE SHOOT APICAL MERISTEM

The shoot apical meristems of adult Echinocereus engelmannii plants are zonate and have a tunica, central mother cells, a peripheral zone, and a pith-rib meristem. An ultrastructural, stereological study showed that each zone has its own distinct ultrastructure, but that the differences between the zones are quite small, both on a protoplasmic basis and on a cytoplasmic basis. Furthermore, the ultrastructure present in the adult apices differed only slightly from that which had been found in seedling apices, demonstrating a long-term stability of structure. The standard deviations found in the sample were small, indicating little variability from one plant to the next and suggesting that there are little or no cyclic changes during the plastochron or a 24-hr photoperiod. The ultrastructures found in the shoot apical meristems differed significantly and markedly from mature tissues of the same plants.     

THE PICEA ABIES SHOOT APICAL MERISTEM IN CULTURE. II. DEPOSITION OF POLYSACCHARIDES AND LIGNIN-LIKE SUBSTANCES BENEATH CULTURES

Excised shoot apical meristems of Picea abies seedlings grow and develop primordial leaves when cultured on Millipore (mixed esters of cellulose) filter membranes lying on a simple, defined medium gelled with agarose. When the cultures are removed from the membranes, each leaves a spot of altered light transmission, spectral characteristics, hygroscopicity, and chemical reactivity. These spots are the manifestation of deposition in the membrane pore space of polysaccharides, lignin-like components, and probably other substances. Deposition of water-insoluble, Schiff's reagent-positive substances can be detected in the filter membranes after only 3–6 hr exposure to a meristem and continues for 10–15 days or longer. Precursors of the insoluble deposition materials can diffuse through at least nine layers of Millipore membrane before deposition at a site remote from living cells. Placement of a dialysis membrane between the meristem and the Millipore membrane prevents detectable deposition in the latter. The observations are consistent with the hypothesis that apical meristems can synthesize and export mobile precursors of cell wall components as well as any substances necessary to promote their condensation or polymerization into insoluble materials at remote sites. The system may be useful in studying synthesis of cell wall components and investigating the functional role of growth regulators in shoot apical development.