AN INVESTIGATION OF THE MORPHOGENETIC MECHANISMS WHICH CONTROL THE DEVELOPMENT OF ZONATION IN SEEDLING SHOOT APICAL MERISTEMS

The development of zonation in the shoot apical meristems of 28 species of cacti was examined. At germination the embryonic apex may have one of three types of organization: 1) tunica/corpus; 2) tunica/central mother cells/corpus; 3) tunica/central mother cells/peripheral zone/pith-rib meristem. Apices of the third type have complete zonation and undergo little or no further structural development. Both of the other types develop the missing zones. First, the apices become mitotically active, and peripheral zone characters develop in the outer corpus. Simultaneously, or slightly later, the central mother cells differentiate if they are not yet present. The final step is the formation of the pith-rib meristem. The sequence of appearance of the zones was constant throughout all species examined, but the time of appearance of only one zone could be correlated with any other morphogenetic process: the development of peripheral zone characteristics in the outer corpus occurs with or before the beginning of leaf production. The development of zonation appears to be independent of apical size, shape, or age, either real age or plastochron age. This has been interpreted to indicate that the metabolic mechanism controlling the development of zonation in shoot apical meristems is largely autonomous and independent of other aspects of morphogenesis occurring in the seedling. Also, the fact that leaf initiation and shoot axis production can both occur before the development of either the central mother cells or the pith-rib meristem indicates that neither of these zones is essential for these two apical morphogenetic activities.     

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.     

THE SHOOT APEX OF BOUGAINVILLEA SPECTABILIS

The shoot apex of Bougainvillea spectabilis consists of five zones: A two- or occasionally three-layered tunica, a central mother cell zone, a cambium-like zone, a rib meristem (central meristem), and a peripheral meristem. The presence of a cambium-like zone is somewhat unusual in the apex of vascular plants, having only been reported for a few taxa. In B. spectabilis the cambium-like zone is consistently present throughout the plastochron and all yearly seasonal periods.     

CYTOKININ- AND GIBBERELLIC ACID-INDUCED EFFECTS ON THE STRUCTURE AND METABOLISM OF SHOOT APICAL MERISTEMS IN OPUNTIA POLYACANTHA (CACTACEAE)

The dormant axillary buds of Opuntia polyacantha can be activated by either cytokinins or gibberellic acid. Under the influence of benzylaminopurine (BAP), the axillary bud meristem increases greatly in size and becomes mitotically active. The primordia produced by the meristem develop as normal photosynthetic leaves. Gibberellic acid (GA) also causes the meristem to become mitotically active, but the meristem does not increase in size. The primordia produced under the influence of GA develop as normal cactus spines. Leaf-producing meristems and spine-producing meristems have the same zonation, despite the differences in size. The meristems are composed of a uniseriate tunica, a central mother cell zone, peripheral zone, and a pith rib meristem. The mitotic activity of each of the zones in the leaf-producing meristem differs significantly from the mitotic activity of the corresponding zones in the spine-producing meristem.     

THE QUIESCENT CENTER IN CULTURED ROOTS OF CONVOLVULUS ARVENSIS L.

Cultured roots of Convolvulus arvensis were incubated in 0.2–0.3 μc/ml methyl-³H-thymidine for different intervals of time. In roots supplied with tritiated thymidine for 12 hr, 14 hr, 48 hr, or 14 hr followed by transfer to fresh medium for 24 hr, autoradiographs prepared of serial, longitudinal sections of the root tips showed the presence of a subterminal quiescent center in the root proper at the distal poles of the central cylinder and cortex. In addition, a zone of unlabelled cells in the columella, distal to the root cap initials, was present. In roots supplied continuously with tritiated thymidine for 64 hr, 96 hr, and 120 hr, the quiescent center was either reduced in size or was not present.     

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.     

ONTOGENY OF THE INFLORESCENCE IN CHENOPODIUM ALBUM

Gifford , Ernest M., Jr ., and Herbert B. Tepper . (U. California, Davis.) Ontogeny of the inflorescence in Chenopodium album. Amer. Jour. Bot. 48(8): 657–667. Illus. 1961.—Chenopodium album, a short-day plant, was induced to flower by subjecting it to successive cycles of 7 hr light and 17 hr darkness. After 4 inductive days, the first macroscopic change is evident in the appearance of precocious axillary bud primordia. After 5–6 days, a primordial inflorescence is visible, and after 7–8 days a terminal flower appears on the main inflorescence axis. The vegetative apex has a biseriate tunica, the cells of which are larger than those of the corpus. The cells of the tunica stain lighter, possess larger nucleoli, and are more vacuolate than cells of the subjacent corpus. After photoinduction, the tunica-corpus organization is maintained, and after 4 short-days, the shoot apex possesses a mantle of 3–4 layers of cells because there are few periclinal divisions in the cells of the outer corpus. The cells of the mantle stain uniformly and are more chromatic than those of the underlying tissue. Mitotic activity was recorded in the upper 40-μ segment of the apex. In the vegetative apex, mitotic activity is greater in the lower portion of the segment. Following photoinduction, mitoses increase throughout the apex until a maximum is reached on the 4th day. Also, the plastochronic interval decreases after photoinduction. Nucleoli of cells of the corpus enlarge following induction until all nucleoli of the apex are nearly equal. Included in the paper are discussions of the general morphological differences between vegetative and flowering shoots.     

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.     

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.     

AN INVESTIGATION OF THE PHYLOGENETIC AND ONTOGENETIC VARIABILITY OF SHOOT APICAL MERISTEMS IN THE CACTACEAE

The sizes, shapes and zonations of the shoot apical meristems of 22 species of cacti were examined. This family was chosen because of its great diversity of habits; the more primitive members are nonsucculent. leafy trees and more advanced members are highly specialized “leaf-less” stem-succulents. By combining these measurements with those already in the literature, a sample of almost 70 species was obtained. Apical meristems range in size from only 80 μm in diam in some species to as much as 1.500 μm in diam in others. The shape ranges from being flat to almost hemispherical. Despite the great range in size and shape of the apical meristems, or the range in the morphologies of the leaves and stems which are produced by the meristems. all apices had the usual zonation: tunica, central mother cells, peripheral zone, and pith-rib meristem. The sizes of each of the zones. expressed either as the number of cells per zone or expressed as a percentage of the whole apex. were highly variable. The variation in apical dimensions and zone sizes occurred both phylogenetically and ontogenetically. and this has been interpreted to indicate that the morphogenetic mechanisms which control apical size and zonation are easily modified, both during the development of individual plants and during the evolution of new species.     

THE ROOT APICAL MERISTEM OF OSMUNDA REGALIS

Roots of the Osmundaceae differ from most ferns in having more than one apical cell. The size of the apical initial group, which includes cells that are considered to be apical cells, varies directly with root diameter in Osmunda regalis L. Mitotic indices were 6.63% for apical cells, 7.45% for the entire apical group, 6.25% for distal derivatives, and 7.15% for developing cortical cells. Cytophotometric measurements of Fuelgen-stained nuclei indicated no endopolyploidy in the populations of cells studied. These findings demonstrate that there is no quiescent center in the roots of O. regalis.     

ONTOGENETIC AND HISTOCHEMICAL CHANGES IN THE VEGETATIVE SHOOT TIP OF CHENOPODIUM ALBUM

Gifford , Ernest M., Jr ., and Herbert B. Tepper . (U. California, Davis.) Ontogenetic and histochemical changes in the vegetative shoot tip of Chenopodium album. Amer. Jour. Bot. 49(8): 902–911. Illus. 1962.—A distinct central zone is present in apices of young, rapidly growing seedlings of Chenopodium album. This zone is clearly evident when the plane of section includes developing leaf buttresses and/or newly formed leaf primordia, but is not visible if the cells associated with these developing foliar organs are not in the plane of section. Cells in the central zone contain markedly lower concentrations of RNA than those of the peripheral zone. Differences in DNA, SH-protein, total protein, and histone concentration of cells in the apex parallel the zonal distribution of RNA, but are much less obvious. As the plant ages and the rate of leaf production decreases, the zonal distribution of the above-mentioned compounds becomes less and less obvious. Cells at sites of leaf initiation in older seedlings still have slightly higher RNA concentrations, but a central zone such as is visible in the young seedlings is no longer evident. The axial tunica cells do, however, contain noticeably lower DNA and histone concentrations than the remaining cells of the shoot apex. Changes occur in the size of nucleoli during ontogeny and the possible relationship between nucleolar size and modifying factors is discussed.     

ROOT APEX STRUCTURE IN EPHEDRA MONOSPERMA AND EPHEDRA CHILENSIS (EPHEDRACEAE)

The root meristem of E. monosperma and E. chilensis possesses a central group of distinctive, large cells. These cells have large nuclei with scattered heterochromatin, proplastids with no starch, small vacuoles, mitochondria, few dictyosomes and endoplasmic reticulum cisternae, and lipid deposits. Over a 24 hr labelling period, the large cells fail to incorporate ³H-thymidine, whereas cells both distal and proximal to this region do. A quiescent center which includes these large cells is present therefore. Both species have an extensive root cap, the length being contributed by mitoses in many tiers of cells distal to the quiescent center. The root cap consists of a columella and peripheral regions. Distinctive amyloplasts, an increase in the number of endoplasmic reticulum cisternae and dictyosomes, large vacuoles, and lipid deposits are characteristic of differentiated columella cells. Peripheral cells elongate, lose most of their starch, and are eventually sloughed from the root.     

MITOTIC ACTIVITY IN THE CAMBIAL ZONE OF PINUS STROBUS

Mitotic activity in the cambial zone of 20-year-old Pinus strobus L. trees was determined quantitatively, using mitotic counts from serial tangential sections of sample pieces. Counts from nine cores 1.6 mm in diameter from each sample piece were averaged and expressed as the number of mitoses per core with the sampling error. Estimation of the number of cambial zone cells per core permitted calculation of mitotic indices. In the fourth internode from the ground the first mitoses were observed on April 15 and the last on September 15. The number of mitoses per sample piece was comparable throughout each internode at any one sampling, but the number was higher in internodes within the crown than in those below it. Mitotic index was not appreciably higher within the crown, but it decreased from 3.7 in spring to 2.0 in fall. An internode in the crown sampled May 11–13 showed afternoon peaks in mitotic activity, but an internode below the crown showed no peaks, nor did other internodes sampled later in the season.     

Structure and Cytogenesis of the Shoot Apex of Syringa oblata var. affinis Lingelsh

The structure, growth and mitotic activity of 211 shoot apices of developing sprouts of Syringa oblata var. affinis Lingelsh. in longitudinal sections and 67 in transverse sections have been studied with the view to understanding the nature of zonation patterns and cytogenesis of the apical meristems during a double plastochron. The external morphology and the anatomical structure of the apices in 4 plastochronic stage-early, middle, late Ⅰ and late Ⅱ stages are described. In the shoot apices examined, especially those at late plastochronic stage, the following zones may be delimited: Zone of tunica initials, zone of corpus initials, peripheral zone and zone of rib meristem. The location and orientation of mitotic figures observed in longisections of the apices in 4 plastochronic stages are plotted in diagrams and the mitotic frequency has been calculated. Information obtained from these investigations reveals that the tunica and corpus inititals constitute an active region of the apex, but their mitotic activity changes periodically within the double plastochron. In the middle plastochronic stage when the apex is at its minimal area and the cells of peripheral zone and rib meristem zone have been completely transformed into constituent parts of foliar primordia and the subjacent tissues of the stem and the pith mother cells respectively, the mitotic frequency of the initials is at its maximium and its intensity of mitotic activity is not much lower than that of any other meristematic zone at any stage. When the apical dome is reformed by the activity of these initials in late plastochronic stages, the mitotic frequency of the initials gradually drops and the region of high mitotic frequency shifts to the flank of the apex, the peripheral zone. Anticlinal divisions are predominant in this zone. On the other hand, those cells directly left behind by the corpus initials, which constitute the rib meristem, are vacuolated and marked by the pre- dominance of transverse divisions. Thus the entire zonation pattern reappears. In the next early plastochronic stage, the mitotic frequency of the tunica and corpus initials drops to its mimimium, but other regions of the apex still maintain a high mitotic frequency. It may be concluded that the tunica and corpus initials form a cytogenerative center of the shoot, and the cytohistological zonation is actually a result of the fact that different regions of apical meristems are different in mitotic activety, different in state of cell differentiation and different in their function in morphogenesis.     

FOLIAR ONTOGENY AND HISTOGENESIS IN MAGNOLIA GRANDIFLORA L. I. APICAL ORGANIZATION AND EARLY DEVELOPMENT

Foliar ontogeny of Magnolia grandiflora was studied to elucidate possible unique features of evergreen leaves and their development. The apex of Magnolia grandiflora is composed of a biseriate or triseriate tunica overlying a central initial zone, a peripheral zone and a pith rib meristem. Leaf primordia are initiated by periclinal divisions on the apical flank of the tunica in its second layer. This initiation and expansion is seasonal just as in related deciduous magnolias. Following leaf initiation, a foliar buttress is formed and the leaf base gradually extends around the apex. As growth continues, separation of the leaf blade primordium from the stipule proceeds by intensified anticlinal divisions in the surface and subsurface layers near the base. Marginal growth begins in the blade primordium when it reaches approximately 200 μm in height and results in the formation of two wing-like extensions, the lamina. This young blade remains in a conduplicately folded position next to the stipule until bud break.     

Physiological regeneration of the digestive parenchyma in Convoluta convoluta and Oxyposthia praedator (Turbellaria,Acoela)

Autoradiography has been applied to two acoel turbellarians, Convoluta convoluta and Oxyposthia praedator, to determine the distribution and fate of proliferative cells. In C. convoluta, mitotic figures and nuclei that labelled with [³H]thymidine could be observed in the peripheral parenchyma but not in the middle zone of the central parenchyma. The time required for regeneration of physiologically competent digestive cells was about 10–15 days. In O. praedator, mitotic figures (in metaphase and telophase) were observed in the peripheral parenchyma while none were found in the epidermis either in untreated animals or after treatment with colchicine. Mitotic figures were found only rarely in the central parenchyma and only in its marginal zone. Autoradiographs of O. praedator demonstrated [³H]thymidine incorporation into both the nuclei and the cytoplasm of peripheral parenchymal cells. In the central parenchyma, no nuclei with primary labelling were observed. The digestive parenchyma of the acoels is regarded as a unique histological system involving both specialized cells of the central parenchyma and stem cells located in the peripheral parenchyma.     

INFLORESCENCE DEVELOPMENT IN BRASSICA CAMPESTRIS L.

Vegetative seedlings of the Ceres strain Brassica campestris L., a quantitative, long-day plant, were induced to flower by exposure to a 16-hr, long-day cycle. Cytohistological and cytohistochemical changes associated with inflorescence development were examined. Developing shoot apices were classified in vegetative, transitional, and reproductive stages. The vegetative apex possessed a biseriate tunica, central zone, peripheral zone and pith-rib meristem. The transitional stage at 48 hr was marked by an increase in size and by a stratification of the upper cell layers of the shoot apex with a concurrent decrease of apical cytohistochemical zonation. The reproductive stage was initiated at 58 hr by periclinal cell divisions in the 3rd and 4th cell layers of the flank region. Cytohistochemical zonation in the vegetative apical meristem was restored in the floral apex. An “intermediate developmental” phase was not observed between the vegetative and reproductive stage.     

ONTOGENY AND PHYLLOTAXIS OF THE TERMINAL VEGETATIVE SHOOTS OF MICHELIA FUSCATA

Tucker Shirley C. (Northwestern U., Evanston, Ill.) Ontogeny and phyllotaxis of the terminal vegetative shoots of Michelia fuscata. Amer. Jour. Bot. 49(7): 722–737. Illus. 1962.—Two patterns of symmetry occur in Michelia fuscata In the lead shoots, leaves arise in a 2/5 spiral arrangement which may be either clockwise or counterclockwise. Other shoots are dorsiventrally organized; these shoots produce leaves in a modified ½ phyllotaxis in which the angle between the 2 files of leaves lies between 100° and 150°, according to the particular branch. Both types of shoot have a zonate apical meristem with a biseriate tunica a central initial zone, and a peripheral zone. The apical configuration of cells does not change appreciably during the plastochron. The flat to low-convex outline of the shoot apex is maintained by initiation of the leaves close to the summit of the apex; the diameter of the meristem diminishes greatly after such an initiation. Leaf inception in the subsurface tunica layer is followed by precocious activity of the marginal meristems which extend the stipular flanges completely around the base of the apical meristem. The stipular margins then fuse laterally and form a hood over the apex. A subapical initial meanwhile is active in the leaf blade, where it persists up to the time the leaf is 2 mm high. The most recent primordium is 300 μ high before another leaf is initiated. The vascular system of the stem is a cylindrical network of leaf traces, with 6–12 traces per leaf. The procambium develops acropetally from preexisting vascular strands in the stem below. Elements of the diverse sclereid system differ in shape in different tissues, according to the availability of intercellular space. Goebel's term “Pendelsymmetrie” is discussed with reference to apical activity in Michelia.     

THE DEVELOPMENT OF THE SPOROCARP OF MARSILEA VESTITA

The tissues of the sporocarp of Marsilea vestita undergo profound changes during development. Early in development, the cells of the peripheral tissues, epidermis, hypodermis and layers of the transitional zone between the hypodermis and more internal tissues contain prominent vacuolar bodies. As development proceeds, these vacuolar bodies disappear. Prominent amyloplasts are found only in the guard cells and in the cells of the transitional zone. Later in development the cells of the hypodermis divide periclinally forming two layers which differentiate as macrosclereids. The cells of the outermost layer of the transitional zone differentiate as osteosclereids. Internally, the cells of the sorophore accumulate large amounts of mucilage in the central vacuoles. The peripheral cytoplasm ultimately degenerates leaving just hygroscopic mucilage. The mucilage carbohydrate contains the sugars, rhamnose and arabinose. In the young sorus, only the spore mother cells and the cells of the indusium contain amyloplasts. By the time of meiosis, there is a massive accumulation of starch in the receptacle, stalk and jacket but not in the tapetum of the sporangia. Late in development, the starch disappears and the mega- and microspores become coated with carbohydrate.