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.     

COMPARATIVE FOLIAR HISTOGENESIS IN ACORUS CALAMUS AND ITS BEARING ON THE PHYLLODE THEORY OF MONOCOTYLEDONOUS LEAVES

A comparative histogenetic investigation of the unifacial foliage leaves of Acorus calamus L. (Araceae; Pothoideae) was initiated for the purposes of: (1) re-evaluating the previous sympodial interpretation of unifacial leaf development; (2) comparing the mode of histogenesis with that of the phyllode of Acacia in a re-examination of the phyllode theory of monocotyledonous leaves; and (3) specifying the histogenetic mechanisms responsible for morphological divergence of the leaf of Acorus from dorsiventral leaves of other Araceae. Leaves in Acorus are initiated in an orthodistichous phyllotaxis from alternate positions on the bilaterally symmetrical apical meristem. During each plastochron the shoot apex proceeds through a regular rhythm of expansion and reduction related to leaf and axillary meristem initiation and regeneration. The shoot apex has a three- to four-layered tunica and subjacent corpus with a distinctive cytohistological zonation evident to varying degrees during all phases of the plastochron. Leaf initiation is by periclinal division in the second through fourth layers of the meristem. Following inception early growth of the leaf primordium is erect, involving apical and intercalary growth in length as well as marginal growth in circumference in the sheathing leaf base. Early maturation of the leaf apex into an attenuated tip marks the end of apical growth, and subsequent growth in length is largely basal and intercalary. Marked radial growth is evident early in development and initially is mediated by a very active adaxial meristem; the median flattening of this leaf is related to accentuated activity of this meristematic zone. Differentiation of the secondary midrib begins along the center of the leaf axis and proceeds in an acropetal direction. Correlated with this centralized zone of tissue specialization is the first appearance of procambium in the center of the leaf axis. Subsequent radial expansion of the flattened upper leaf zone is bidirectional, proceeding by intercalary meristematic activity at both sides of the central midrib. Procambial differentiation is continuous and acropetal, and provascular strands are initiated in pairs in both sides of the primordium from derivatives of intercalary meristems in the abaxial and adaxial wings of the leaf. Comparative investigation of foliar histogenesis in different populations of Acorus from Wisconsin and Iowa reveals different degrees of apical and adaxial meristematic activity in primordia of these two collections: leaves with marked adaxial growth exhibit delayed and reduced expression of apical growth, whereas primordia with marked apical growth show, correspondingly, reduced adaxial meristematic activity at equivalent stages of development. Such variations in leaf histogenesis are correlated with marked differences in adult leaf anatomy in the respective populations and explain the reasons for the sympodial interpretation of leaf morphogenesis in Acorus and unifacial organs of other genera by previous investigators. It is concluded that leaf development in Acorus resembles that of the Acacia phyllode, thereby confirming from a developmental viewpoint the homology of these organs. Comparison of development with leaves of other Araceae indicates that the modified form of the leaf of Acorus originates through the accentuation of adaxial and abaxial meristematic activity which is expressed only slightly in the more conventional dorsiventral leaf types in the family.     

A SURVEY OF THE VEGETATIVE SHOOT APICES IN THE FAMILY MALVACEAE

The present study compares the structure of the vegetative shoot apex in 40 species of the Malvaceae. There is a wide range of size, shape, and zonation within the apices of the family. Although many of the apices are domed, some are flat-topped and do not extend above the axil of the youngest leaf primordium. Also, most of the species investigated are recorded as having a more or less marked cytohistological zonation superimposed on the tunica-corpus configuration. The tunica is single-layered in a majority of species, but stratification of the upper corpus is common. In an effort to give a more accurate concept of apical structure and activity, the apex is described as the metrameristem and its derivatives: the flanking meristem, and the pith rib meristem or pith mother cells. The metrameristem, consisting of the tunica initials and the co pus initials, is the focal point of the study of the zoned apices. Data are presented for the measurements of the metrameristem, measurements of the apical dome, type of flanking meristem, origin of the pith, and growth habit of the plant. There appears to be a correlation between growth habit and the distinctness with which the metrameristem is marked off from the surrounding tissue. Most of the herbaceous species have an indistinctly marked metrameristem, whereas the shrubby trees and trees have a distinctly marked metrameristem. Zonation in shrubs and suffrutescent plants may be of either type.     

THE SHOOT APEX AND EARLY LEAF DEVELOPMENT IN CLEMATIS

Tepfer , Sanford S. (U. Oregon, Eugene.) The shoot apex and early leaf development in Clematis . Amer. Jour. Bot. 47 (8): 655–664. Illus. 1960.—The high-domed shoot apex comprises a 2-layered tunica and shallow corpus. The rib meristem at times extends to within 5 cells of the summit. The cells of tunica and corpus are uniform cytologically, distinguishable only by the orientation of division planes. No zonation is visible within the corpus. No evidence was found of the existence of a méristème d'attente; mitotic figures appear frequently in the central region of the tunica and corpus. Decussately arranged leaf primordia arise high on the flanks of the apex. Periclinal divisions in the inner tunica and outermost corpus layers mark the site of initiation. Details of the growth and early differentiation of the leaf primordia follow the usual pattern of buttress formation, growth through apical and subapical initials. Apical growth continues beyond the early stages of leaf ontogeny; the blade-forming marginal meristems do not appear until after leaflet primordia are formed. There are 5 primary leaflets, pinnately arranged. Each leaflet is 3- to 5-lobed. In primordium P₃ expansion of the adaxial-lateral margins occurs at the base, but not above. This marks the upper limits of the basal pair of lateral leaflets. In P₄ the upper limits of the upper lateral leaflets become demarcated in similar fashion.     

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.     

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.     

HISTOGENESIS OF THE ANDROECIUM AND GYNOECIUM IN DOWNINGIA BACIGALUPII

A histogenetic investigation of the synandrous androecium and syncarpous gynoecium in the flower of Downingia bacigalupii Weiler (Campanulaceae; Lobelioideae) was undertaken for the purpose of comparing the modes of initiation, early growth and fusion in these floral whorls with that reported previously for the perianth in this species. Stamens are initiated as separate organs from the second tunica layer and underlying corpus regions of the concave floral meristem. Subsequent growth of stamens involves apical and intercalary growth in length and rudimentary marginal growth in breadth. Tissues of the four microsporangia originate from hypodermal sporangial initial cells and the filament is formed by intercalary growth at the base of the anther. Lateral fusion of stamens is ontogenetic and involves cuticular fusion of adjacent epidermal layers. The two emergent carpel primordia arise as crescentic organs by periclinal divisions in the second tunica layer and corpus zones. Carpel primordia also undergo apical and intercalary growth in length as well as extensive marginal growth in breadth. Radial growth in carpels is mediated by an adaxial meristem which shows its greatest concentration of activity at the carpel margins. Carpel fusion appears to be partially ontogenetic accompanied by zonal growth. Closure of the stylar canal is by the formation of a transmitting tissue derived from the protodermal layers of the adaxial carpel surfaces. A discoid nectary is initiated around the base of the style and formation of the inferior ovary is by intercalary growth of the base of the concave floral bud. The two parietal placentae originate as longitudinal outgrowths from the walls of the floral cup. Ovule initiation is simultaneous at first and then intercalary during subsequent elongation of the ovary. The ovules are anatropous, unitegmic and tenuinucellate. Stamen and carpel procambium shows a slight delay in differentiation when compared to that reported for the perianth and bract, but in all other respects carpels resemble other floral organs in their patterns of histogenesis and early growth. Stamens diverge from the other floral organs in their early pattern of growth, but a consideration of all features of their histogenesis suggests an appendicular rather than an axial interpretation of these organs.     

The Stem Cell Niche in Leaf Axils Is Established by Auxin and Cytokinin in Arabidopsis

Plants differ from most animals in their ability to initiate new cycles of growth and development, which relies on the establishment and activity of branch meristems harboring new stem cell niches. In seed plants, this is achieved by axillary meristems, which are established in the axil of each leaf base and develop into lateral branches. Here, we describe the initial processes of Arabidopsis thaliana axillary meristem initiation. Using reporter gene expression analysis, we find that axillary meristems initiate from leaf axil cells with low auxin through stereotypical stages. Consistent with this, ectopic overproduction of auxin in the leaf axil efficiently inhibits axillary meristem initiation. Furthermore, our results demonstrate that auxin efflux is required for the leaf axil auxin minimum and axillary meristem initiation. After lowering of auxin levels, a subsequent cytokinin signaling pulse is observed prior to axillary meristem initiation. Genetic analysis suggests that cytokinin perception and signaling are both required for axillary meristem initiation. Finally, we show that cytokinin overproduction in the leaf axil partially rescue axillary meristem initiation-deficient mutants. These results define a mechanistic framework for understanding axillary meristem initiation.     

CONSTANCY OF RELATIVE VOLUMES OF ZONES IN SHOOT APICAL MERISTEMS IN CACTACEAE: IMPLICATIONS CONCERNING MERISTEM SIZE,SHAPE, AND METABOLISM

The volumes and dimensions of shoot apical meristem zones were determined for 21 species in 10 genera of the Cactaceae. If the volumes of the central-mother-cell zone (CMCZ), the peripheral zone (PZ), and the pith-rib meristem (PRM) are expressed as percentages of the total volume of the corpus, then they are remarkably constant (CMCZ = 11.1% of the volume of the corpus, PZ = 61.2% and PRM = 27.1%) regardless of the relative size of the apical meristems. The relative volume of the tunica decreases, however, as the whole apex increases. The zones have variable shapes, and whereas the PZ and PRM are always trapezoidal in longitudinal section, in apices with large volumes, these zones have a flatter, more flaring shape than they have in smaller apices. Thus large apices are flatter and less hemispherical than are small apices. The CMCZ, however, maintains a constant shape and is always relatively hemispherical regardless of its volume. A hypothesis that relates all of these shape and volume relationships as an integrated growth sequence is proposed.     

Activity and accumulation of cell division-promoting phenolics in tobacco tissue cultures

Dehydrodiconiferyl alcohol glucosides (DCGs) are derivatives of the phenylpropanoid pathway that have been isolated from Catharansus roseus L. (Vinca rosea) crown gall tumors. Fractions containing purified DCGs have been shown previously to promote the growth of cytokinin-requiring tissues of tobacco in the absence of exogenous cytokinins. In this study, we utilized synthetic DCG isomers to confirm the cell division-promoting activity of DCG isomers A and B and show that they neither promote shoot meristem initiation on Nicotiana tabacum L., cv Havana 425, leaf explants nor induce betacyanin synthesis in amaranth seedlings. Analysis of cultured tobacco pith tissue demonstrated that DCG accumulation was stimulated by cytokinin treatment and correlated with cytokinin-induced cell division. Thus, the accumulation of metabolites that could replace cytokinin in cell division bioassays is stimulated by cytokinins. These data support the model that DCGs are a component of a cytokinin-mediated regulatory circuit controlling cell division.     

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.     

THE EARLY ACTION OF THE FLORAL STIMULUS ON MITOTIC ACTIVITY AND DNA SYNTHESIS IN THE APICAL MERISTEM OF XANTHIUM STRUMARIUM

Vegetative plants of Xanthium strumarium (a short-day species) were induced to flower by exposure to a single 16-hr long night. By cutting off the induced leaf (half-expanded leaf) at various times, it was established that, by 8 hr after the end of the long night, a sufficient amount of floral stimulus had reached the meristem to induce a flowering response. The following sequence of events occurred in both the peripheral and central zones of the apical meristem of induced plants: 1) a rise in the mitotic index beginning at 28 hr after the end of the long night and culminating at 36 and 56 hr; 2) a stimulation of DNA synthesis starting at 32–36 hr and reaching a maximum at 60 hr; 3) an increase in nucleolus diameter starting at 32 hr. The cell population in the meristems of both vegetative and induced plants displayed a similar distribution, with about 80 % of the nuclei with the 2C amount of DNA. The comparison of the kinetic data concerning the mitotic index and DNA synthesis indicated that one of the early effects of the floral stimulus in the peripheral and central zones was the release in mitosis of cells whose nuclei were in the postsynthetic (G₂) phase of the mitotic cycle. In the pith-rib meristem, the following events were recorded: 1) a stimulation of DNA synthesis starting at 20 hr; 2) a rise of the mitotic index beginning at 28 hr; 3) the vacuolation and elongation of cells starting at 48 hr. All these events occurred well before the initiation of bract and flower primordia, which began at 96 and 136 hr, respectively. Neither stimulation of mitotic activity nor flowering occurred in the meristems of plants subjected to a long night interrupted at its midpoint by a 5-min light break. The results are discussed in relation to the early events which are known to occur in the meristems of other photoperiodic species in transition to flowering.     

HISTOCHEMICAL STUDY OF ENZYME ACTIVITY IN THE SHOOT APICAL MERISTEM OF BRASSICA CAMPESTRIS DURING TRANSITION TO FLOWERING. IV. GLUCOSE-6-PHOSPHATASE

Glucose-6-phosphatase (G6P) activity was determined in fresh-frozen, cryostat sections in the shoot apical meristem of Brassica campestris L. Enzymatic activity was differentially distributed in a zonate pattern in the vegetative meristem, but not in the transition and floral meristem. Vegetative apices showed a heterogenous localization with the highest activity in the central zone and the pith-rib meristem zone. At the early transition stage of development, G6P activity in the peripheral zone increased slightly. At the late transitional (prefloral) stage, G6P activity was not localized within the peripheral zone in island-like areas of activity. This is the first demonstration of G6P in shoot apical meristem at the vegetative, transition, and floral stage. The results indicate that G6P activity 1) is an accompanying event of evocation, but 2) does not mark incipient floral primordia. G6P may play an important role in the maintenance of glucose-6-phosphate homeostasis in an evoked shoot apical meristem.     

Characterization of an Arabidopsis gene that mediates cytokinin signaling in shoot apical meristem development

Cytokinins are adenine derivatives that regulate numerous plant growth and developmental processes, including apical and floral meristem development, stem growth, leaf senescence, apical dominance, and stress tolerance. However, not much is known about how cytokinin biosynthesis and metabolism is regulated. We identified a novel Arabidopsis gene, ALL, encoding an aldolase-like enzyme that regulates cytokinin signaling. An Arabidopsis mutant, all-1D, in which ALL is activated by the nearby insertion of the 35S enhancer, exhibited extreme dwarfism with rolled, dark-green leaves and reduced apical dominance, symptomatic of cytokinin-overproducing mutants. Consistent with this, ARR4 and ARR5, two representative primary cytokinin-responsive genes, were significantly induced in all-1D. Whereas SHOOT MERISTEMLESS (STM) and KNAT1, which regulate meristem development, were also greatly induced, expression of REV and PHV that regulate lateral organ polarity was inhibited. ALL encodes an aldolase-like enzyme that belongs to the HpcH/HpaI aldolase family in prokaryotes and is down-regulated by exogenous cytokinin, possibly through a negative feedback pathway. We propose that ALL is involved in cytokinin biosynthesis or metabolism and acts as a positive regulator of cytokinin signaling during shoot apical meristem development and determination of lateral organ polarity.     

Effect of thidiazuron,light fluence rates and kanamycin on in vitro shoot organogenesis from excised Rubus cotyledons and leaves

The relationship between genotype, tissue age and endogenous cytokinin levels on adventitious bud formation on Lachenalia leaf tissue were investigated. The genotypes studied, showed a variation in bud formation. The hybrid explants responded differently to factorial combinations of BA and NAA. The growth regulators could not substitute for the regeneration potential of the genotype. Tissue age had a pronounced effect on regeneration potential. Young tissue formed the largest number of buds. An interaction between tissue age and genotype was detected. Cytokinin levels in young leaf tissue were higher than in older tissue. In young tissue no relationship was observed between the cytokinin level and the number of buds formed. However, in older tissue it appears as if a relatively low endogenous cytokinin level enhanced bud formation.Abbreviations BA benzyladenine - NAA naphthalene-1-acetic acid - Z zeatin - ZR ribosylzeatin     

OBSERVATIONS ON LEAF AND BUD FORMATION IN HYDROCHARIS MORSUS-RANAE

Regular sequences of leaf and bud formation occur in several members of the Hydrocharitaceae, including Hydrocharis, in which buds are normally formed in the axil of every second leaf of the phyllotactic spiral. Leaf inception begins by periclinal divisions of the inner cells of the 2-layered tunica. Bud formation, which occurs in the apical meristem itself, immediately following the inception of the subtending leaf primordium, begins by divisions in various planes in the corpus, the 2 tunica layers remaining continuous throughout. The young bud meristem soon gives rise to a lateral bud, before leaf formation begins upon it. Because of these and other features, this species is one of considerable morphogenetic interest. Morphogenesis of the whole plant, and in particular the factors controlling the regular sequence of leaf and bud formation, have been and are being investigated experimentally.