Glucose-6-phosphate-dehydrogenase activity in the shoot apical meristem,leaf primordia,and leaf tissues of Dianthus chinensis L.

The oxidation of carbohydrate by the pentose-phosphate pathway in the shoot apical meristem and developing leaf primordia of Dianthus chinensis was assessed by measuring the activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49). On a kg^-1 dry weight h^-1 basis, activity rose from 250 mmol in the apical meristem to 550 mmol in the first two leaf primordia and then declined to 350 mmol in the sixth pair of leaf primordia, and finally to 200 mmol in leaves just emerged from the shoot bud. Measurements of activity in the sixth leaf pair from the apex showed differential distribution in leaf tissues. Epidermal and mesophyll tissue had about the same activity as whole-leaf tissue, but vascular bundles had 70% greater activity. Within the vascular tissue, activity in the phloem was twice as high as in the xylem. When activity was expressed on a per-cell basis, there was a continuous increase from 20 fmol in the apex to 2 pmol in the sixth leaf pair. Activity on a per unit cell volume basis showed that cells of the apical meristem and the epidermis, mesophyll and xylem of the sixth leaf pair had similar values, about 30 amol; only the two youngest pairs of primordia and the phloem had values two or three times this amount.     

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.     

Latent root primordia in poplar stems

Root primordia initiate in poplar stems in the secondary growing parts, that is in the parts where the elongation growth is terminated and the leaves are mature. Their initiation is connected with the occurrence of unusual biseriate, rarely multiseriate rays. A small cell group in the secondary phloem is initiated by cell division of the ray. It gradually enlarges by continuing cell division, by the addition of cells adjacent to the cell group and by cambial activity. Thus, a hemispherical root primordium is formed, for which a permanent occurrence of reserve lipids is characteristic. In stems several years old the intraprimordial mitotic activity is rhythmically renewed together with the cambium function renewal. Latent root primordia slightly enlarge with the passing years, whereas mainly the cells localized in their centre divide. Further organization and root histogenesis was not observed either in older root primordia. Adjacent to root primordia, cambial initials produce the secondary xylem elements increasingly. Xylem protuberances are thus formed under root primordia. Primordia initiation is most frequent within the first year of stem development, though they can also initiate in later years.     

CYTOKININ- AND GIBBERELLIC ACID-INDUCED EFFECTS ON THE DETERMINATION AND MORPHOGENESIS OF LEAF PRIMORDIA IN OPUNTIA POLYACANTHA (CACTACEAE)

Cytokinins and gibberellins are able to strongly influence the development of “leaf” primordia in the cactus Opuntia polyacantha. Under the influence of cytokinin, the primordia produced by cultured axillary bud apical meristems develop as normal, photosynthetic leaves, being composed of regular epidermal cells, guard cells, mesophyll and mucilage cells as well as vascular tissue. Under the influence of gibberellic acid (GA), the primordia develop as cactus spines, composed of thick-walled epidermal and fiber cells. Guard cells, vascular tissue and parenchyma do not occur. Thus GA is able to redirect leaf morphogenesis in O. polyacantha far more completely than has been reported for other plants. The mitotic activity of the primordia that will develop into spines is significantly higher (at the 5 % level) than the mitotic activity of the primordia that will develop into leaves. This is interpreted to indicate that the primordia are either leaf primordia or spine primordia from a very early age, and possibly are never uncommitted or undetermined primordia, as has been suggested for fern leaf primordia.     

The Response of Primordial Cells of Vicia faba to Colchicine

Whole root systems of Vicia faba were treated with 0.025 percent colchicine for 3 h. Meristems of roots and developing primordiawere examined immediately after treatment and 24 and 48 h later.Large primordia, i.e. those primordia that would emerge as lateralroots within approximately 24 h, are induced to form tetraploidcells. However, unlike lateral roots, they do not form a typicalc-tumour, their longitudinal growth is not inhibited, and theirmitotic index does not fall. Furthermore, the tetraploid cellsinduced in large primordia are not maintained when the primordiaemerge as lateral roots. Since the response of the very largestprimordia to a 3-h treatment with 0.025 per cent colchicineis similar in every way to that of lateral roots, it appearsthat the cells of primordia acquire the characteristics of lateralroot meristematic cells before the primordia emerge. From theevidence that induced tetraploid cells fail to survive as aprimordium grows out to form a lateral, it also seems that astructural reorganization accompanies the physiological reorganizationthat occurs as a primordium emerges.     

Developmental Morphology and Histogenesis of Reproductive Structures of the Millet,Panicum miliaceum L. Histogenesis of the Inflorescence and Flower

The branches of successive orders of the inflorescence of Panicum miliaceum L. arise in the axils of the bracts of the branches of next lower order. Their initiation is evidenced by periclinal division of sub-hypodermal cells. The primordia of branches arise in initiation like a normal axillary bud. The floral histogenesis of Panicum miliaceum L. is similar to that of Triticum. Primordia of the spikelet, flower and stamen are initiated by the activity of the periclinal division of the sub-hypodermal cell or cells. Sometimes, periclinal divisions also occur in a few hypodermal cells during these primordial developments; such divisions are more frequent in the formation of the flower and stamen primordia than in the formation of the spikelet primordia. The periclinal division of the dermatogen ceils never occurs in the formation of these organs. Glumes and lemma are initiated in the periclinal division of the dermatogen and hypodermal cell or cells. The primordia of the palea, lodicule and carpel are initiated by means of the periclinal division in the dermatogen cell or cells. In the formation of the palea and carpel, periclinal divisions also occur in hypodermat cells, but their derivatives are protruding into the bases of the primordia and do not constitute the tissues of the palea and carpel. The growing point of the flower axis develops into the ovule. The integuments arise from the periclinal division of dermatogen cells. The periclinal division of dermatogen cells is characteristic of the initiation of the phylloid organs in the Gramineae.     

Key proliferative activity in the junction between the leaf blade and leaf petiole of Arabidopsis

Leaves are the most important, fundamental units of organogenesis in plants. Although the basic form of a leaf is clearly divided into the leaf blade and leaf petiole, no study has yet revealed how these are differentiated from a leaf primordium. We analyzed the spatiotemporal pattern of mitotic activity in leaf primordia of Arabidopsis (Arabidopsis thaliana) in detail using molecular markers in combination with clonal analysis. We found that the proliferative zone is established after a short interval following the occurrence of a rod-shaped early leaf primordium; it is separated spatially from the shoot apical meristem and seen at the junction region between the leaf blade and leaf petiole and produces both leaf-blade and leaf-petiole cells. This proliferative region in leaf primordia is marked by activity of the ANGUSTIFOLIA3 (AN3) promoter as a whole and seems to be differentiated into several spatial compartments: activities of the CYCLIN D4;2 promoter and SPATULA enhancer mark parts of it specifically. Detailed analyses of the an3 and blade-on-petiole mutations further support the idea that organogenesis of the leaf blade and leaf petiole is critically dependent on the correct spatial regulation of the proliferative region of leaf primordia. Thus, the proliferative zone of leaf primordia is spatially differentiated and supplies both the leaf-blade and leaf-petiole cells.     

Meristematic Activity during Adventitious Root Primordium Development: Influences of Endogenous Auxin and Applied Gibberellic Acid

Intact brittle willows (Salix fragilis L.) were treated so that developing adventitious root primordia in the stems would be subjected to elevated gibberellic acid or reduced endogenous auxin levels. Observations were made of primordia that were initiated during the experiments and of primordia that were established before the experiments began. The results indicated that as primordia became older and contained more cells, auxin basipetally transported in the stem seemed to be of less importance in determining cell number per primordium. Thus, established primordia depended upon this auxin to a lesser extent than primordia which were being initiated. These observations were explained on the basis of differential contributions during primordium development of cell division in the cambium of the stem and in the primordia themselves. As opposed to the effects of reduced auxin levels, applied gibberellic acid reduced the cell number per primordium most in established primordia. Initiating primordia were least affected by gibberellic acid treatment. Gibberellic acid treatment seemed mainly to reduce intraprimordium cell division, on which continued development of established primordia most depends. Seemingly, at least in brittle willow, applied gibberellic acid blocks the action of auxin in primordium development subsequent to the initiation phase.     

Early development of leg and wing primordia in the Drosophila embryo

The development of the leg and wing primordia in the Drosophila embryo has been traced using molecular markers. Distal-less and disconnected gene expression provide molecular labels for the leg primordia throughout embryonic development, disconnected expression in the developing leg primordia depends on Distal-less activity. The leg primordia arise as discrete clusters of cells that occupy well defined positions in the embryonic ectoderm. At later stages of embryogenesis the primordia become morphologically recognizable and are intimately associated with the development of the Keilin's organs. The presumptive leg disc and the Keilin's organ appear to derive from a common primordium. Similarly the Abnormal leg pattern gene provides a molecular label for the wing and haltere primordia. The dorsal thoracic primordia appear to be of independent origin from the legs.     

Coiled bodies in nuclei from plant cells evolving from dormancy to proliferation

To characterise the coiled bodies in meristematic nuclei of Saccharum officinarum, immunofluorescence labelling with antibodies against components of the splicing (U2B' and Sm core protein B) and pre-rRNA processing (fibrillarin) complexes was used in cells from the dormant root primordia and from roots at different times after activation to the steady state of proliferation. The number, size and distribution of coiled bodies varied in the meristematic tissue depending on cell activity. While G0 cells in the dry primordia and proliferating cells showed a similar number of coiled bodies attached to their nucleoli, the number of nucleoplasmic coiled bodies greatly increased after the primordia were stimulated to proliferate. Their number remained steady from the time the meristematic population reached the steady state of proliferation, as estimated by flow cytometry. Fractionation studies demonstrated that coiled bodies are a part of the underlying nuclear matrix. Comparison of immunocytochemical and cytochemical data from confocal and electron microscopical studies demonstrated that the nucleolar and nucleoplasmic coiled bodies detected by confocal microscopy shared many features, suggesting that they form a family of closely related structures.     

Studies on induced changes in growth patterns and polarity in roots of Vicia faba L.

Summary Roots of Vicia faba were treated with colchicine (0.025%), or IAA (4.7×10^-6 M), or both, for 3 hours and fixed at various intervals over the following 11 days. The axis of spindle orientation and the distribution of mitotic figures, lateral root primordia and xylem vessel elements was examined in the apical 10 mm of median longitudinal sections of these roots.No effect of IAA was found on the orientation of the spindle. However, evidence was obtained indicating that the systems controlling the polarity of cell division and cell expansion differ in some way.The number of lateral root primordia formed was greater in roots treated with IAA or colchicine than in control roots. These primordia were always initiated adjacent to a xylem vessel. Thus, no primordium was closer to the apex than the most apical xylem vessel, suggesting that an endogenous factor involved in primordia initiation is transported in the xylem. The primordia which develop after colchicine treatment grow out as lateral roots; this is in contrast with those which form after IAA treatment and which do not undergo elongation. These results, which it must be emphasized apply only to the apical 1 cm of treated roots, indicate that lateral root primordia become sensitive to IAA at a certain stage in their development. Exogenous IAA acts as an inhibitor.The new meristem, which forms in the primary root apex after colchicine treatment, contains both diploid and polyploid cells, i.e. it was formed from cells that were unaffected and from cells that were affected by colchicine. Following colchicine treatment the size of the meristem shrinks and this can be prevented by treatment with IAA. This and other evidence presented here, suggests that IAA is a factor involved in the control of the size of the apical meristem in normal roots.     

Cell Population Studies in Developing Root Primordia

The proliferation of polyploid and diploid cells was followedduring the development of lateral root primordia of Vicia faba.Presumptive primordial cells were treated with weak solutionsof colchicine, 0.003, 0.006, and 0.0125 per cent, for 3 h. Somecells are induced to become polyploid and they form a markedsub-population. The relative frequencies of diploid and polyploidcells were determined from the time small primordia were presenttill lateral roots were fully emerged. These frequencies havebeen found to change in a regular fashion and the changes reflectdifferent levels of mitotic activity in different parts of aprimordia as it develops. Polyploid cells occupy the centralcore of the primordia and they divide actively except for theperiod just before lateral root emergence. Coincident with thistemporary quiescence of the core cells, the peripheral cells,which remain diploid after treatment with the weaker solutionsof colchicine, continue to divide. These changes in mitoticactivity of the central and peripheral cells are shown bothin the changes in the relative frequencies of polyploid anddiploid mitoses as a primordium grows and in changes in mitoticindex. A microspectrophotometric study indicates that the centralcells are held, temporarily in the G₁ phase of the cell cycle.     

Development of the adult leg epidermis in <Emphasis Type="Italic">Manduca sexta</Emphasis>: contribution of different larval cell populations

During metamorphosis of the tobacco hornworm Manduca sexta, the simple thoracic legs of the larva are remodeled into the more complex adult legs. Most of the adult leg epidermis derives from the adult primordia, small sets of epidermal cells located in specific regions of the larval leg, which proliferate rapidly in the final larval instar. In contrast, the contribution of the epidermal cells outside the primordia is unknown. In this study we have determined their contribution to the adult leg by labeling them with 5-bromodeoxyuridine (BUdR) and following their fate. Although the labeled cells diminished drastically in number, small groups of these cells persisted into the midpupal stage suggesting that they do contribute to the adult leg epidermis. We also found that during the wandering stage the adult primordia went through active proliferation and very little cell death, while the cells outside the primordia went through extensive cell death accounting for the decrease in their number. Our results indicate that two distinct cell populations exist outside the adult primordia. Most cells belong to the first population, which is larval-specific and disappears through apoptosis early in metamorphosis. The second population consists of polymorphic cells that contribute to the larval, pupal and adult leg epidermis.Edited by D. Tautz     

A Differential Response to Colchicine of Meristems of Roots of Vicia faba

In whole root systems of Vicia faba, including primaries andlaterals, the meristems do not respond in a uniform manner totreatment with colchicine. The meristems of the primary andfully emerged lateral roots become mixoploid and these rootsshow a temporary inhibition of growth. Lateral roots that emergewithin 48 hours of treatment show no effects of colchicine;their growth is not inhibited and they contain few or no polyploidcells. The cells of the primordia that produce these lateralsappear to be insensitive to colchicine: at the insensitive stagethese primordia contain at least 1, 000 cells. Primordia withless than 800 cells are very sensitive to colchicine. They donot recover from treatment and appear to be completely inhibited.The stages of extreme sensitivity and resistance are transientphases in the morphogenesis of a lateral root. The change insensitivity to colchicine is accompanied by a fall in the mitoticindex; this is highest in young primordia, where it is abouttwice the value found in growing laterals.     

A molecular mechanism that confines the activity pattern of miR165 in Arabidopsis leaf primordia

In Arabidopsis leaf primordia, the expression of HD‐Zip III, which promotes tissue differentiation on the adaxial side of the leaf primordia, is repressed by miRNA165/166 (miR165/166). Small RNAs, including miRNAs, can move from cell to cell. In this study, HD‐Zip III expression was strikingly repressed by miR165/166 in the epidermis and parenchyma cells on the abaxial side of the leaf primordia compared with those on the adaxial side. We also found that the MIR165A locus, which was expressed in the abaxial epidermis, was sufficient to establish the rigid repression pattern of HD‐Zip III expression in the leaf primordia. Ectopic expression analyses of MIR165A showed that the abaxial‐biased miR165 activity in the leaf primordia was formed neither by a polarized distribution of factors affecting miR165 activity nor by a physical boundary inhibiting the cell‐to‐cell movement of miRNA between the adaxial and abaxial sides. We revealed that cis‐acting factors, including the promoter, backbone, and mature miRNA sequence of MIR165A, are necessary for the abaxial‐biased activity of miR165 in the leaf primordia. We also found that the abaxial‐determining genes YABBYs are trans‐acting factors that are necessary for the miR165 activity pattern, resulting in the rigid determination of the adaxial–abaxial boundary in leaf primordia. Thus, we proposed a molecular mechanism in which the abaxial‐biased patterning of miR165 activity is confined.     

Hypothalamic influence on differentiation of the immunoreactive ACTH beta-LPH, alpha- and beta-endorphin containing cells in adenohypophysial primordia of rat fetus in organ culture (author's transl)]

Adenohypophysial primordia were isolated in rat fetuses from day 12.5 to day 15.5 of gestation. The organ culture employed for maintenance of the primordia was made up according to Watanabe et al. (1973). The fixation of primordia in Bouin Hollande's solution was performed after 9, 8, 7 or 6 days of culture when the normal duration of pregnancy was achieved. The cultivated primordia were immunologically studied using different antisera: anti-alpha(17-39)ACTH, anti-beta(1-24)ACTH, anti-beta-LPH, anti-alpha and anti-beta-endorphins, with immunoperoxidase or immunofluorescence techniques, including control experiments of the specificity of the antisera. A similar study was performed on pituitaries removed from normal rat fetuses from day 16.5 of gestation and each day up to birth, and fixated immediately. In vivo the first cells reacting with all the antisera used in this study were observed on day 16.5 of gestation; their number increased during gestation (Fig. 1 A, B and C). Immunoreactive cells with the different antisera could be detected in primordia isolated on day 12.5 of gestation after 9 days of culture. Numerous groups of cells were observed in primordia of older fetuses (Fig. 2 A and B). These data indicate that the corticotropic cells in rat fetuses could start to be differentiated without stimuli from the hypothalamus since primordia were isolated before the appearance of this cell type in normal rat fetuses and before the differentiation of the hypothalamus. The presence of ACTH and other peptides such as beta-LPH or beta-endorphin would support the hypothesis of a common precursor in this cell type existing early in gestation. Similar results were obtained in human fetuses.     

Growth and Development of the Banana Plant: II. The Transition from the Vegetative to the Floral Shoot in Musa acuminata cv. Gros Michel

The changes that occur in the shoot apex of the banana, as itpasses from the vegetative to the flowering stage, are described.The crucial events occur well before floral primordia are evident,and they require a redistribution of activity in the variousgrowing regions. The vegetative shoot apex is in a central depressionin the rhizome; there is virtually no internodal growth in theaxis, the most active growth is in the leaf bases; vegetativebuds do not form in the leaf axils but only appear adventitiouslyfar from the tip of the shoot. With the onset of flowering thisis changed; growth in the axis itself, previously suppressed,occurs and flower buds arise as primordia in the axils of subtendingbracts. The bracts do not show the market growth in their baseswhich is so characteristic of leaves. Thus, the shoot apex risesto the level of the rhizome and then above it; as it does so,its tip changes in shape from a broad flattened some to a pointedcone. At the transitional stage, more activity occurs in thecells of the mantle, or tunica, which now consists of 3 to 4layers over the central dome. Below, in the central or mothercell zone of the corpus, which was quiescent in the vegetativeshoot, the cells spring into greater activity, becoming moreprotoplasmic and stain more deeply. Directly below this regionin the rib meristem, cells show transverse divisions. Bractprimordia occur high on the flanks of the apex, and, thoughthey originate in the manner of leaves, their subsequent growthis different. Flower primordia occur even in the axils of bractsclose to the shoot tip. Thus, the problem now is to designatethe source, nature, and mode of action of the stimuli whichinitiate and control this quite different distribution of growthin the floral, as contrasted with the vegetative, shoot. Thesignificance of the previously more quiescent central, or mothercell zone, of the apex as the source of such stimuli, is stressed.Thus, flowering first requires that the limiting controls whichapply to the vegetative shoot be released, and, secondly, thatthe apex of the shoot, rather than the leaf base, becomes themain centre of growth and development.     

Histochemical investigation of β-glucuronidase in culture cells and regenerated plants of Scutellaria baicalensis Georgi

 Strong activity of β-glucuronidase first appeared in the epidermal and glandular hair cells of leaf primordia regenerated from callus of Scutellaria baicalensis Georgi. Leaf primordia matured rapidly in culture to form shoots within 1 month in which both the mesophyll cells and the glandular hairs were deeply stained. Leaves predominantly accumulated β-glucuronidase in both glandular hair cells and mesophyll cells. β-Glucronidase activity in leaves was higher in the summer and decreased in the winter. The stem section collected in the summer had a different β-glucuronidase distribution pattern from that of the root in that in the former strong activity appeared in the periderm cells and collenchyma cells which was decreasingly dispersed into the phloem layer cells. In the winter, β-glucronidase activity decreased compared to that in summer. It can be argued that the distribution of β-glucuronidase in this plant is closely linked with the defense against pathogens: it is a starting key enzyme which may act together with the flavonoids, which play an important role as a proton donor for the detoxification metabolism of H₂O₂. Received: 1 December 1998 / Revision received: 10 March 1999 / Accepted: 14 June 1999     

Mitotic activity in wheat shoot apical meristems: effect of dissection to expose the apical dome

An efficient method, less laborious than histological procedures, is described to screen relatively large numbers of shoot apices for mitotic activity. Mitotic activity of shoot apices of Triticum aestivum L. was observed by differential interference contrast (DIC) microscopy of apices infiltrated with a clearing fluid (chloral hydrate/phenol/lactic acid/dibutylphthalate/benzyl benzoate). Serial optical sections were viewed through entire vegetative apical domes and floral primordia. In vegetative shoots, mitotic cells were observed throughout the light and dark cycles of plants maintained in either 8 or 16 h photoperiods. Mitotic activity was lower in the dark phase and increased through the light cycle in both photoperiods. Cells in L1 and L2 layers at the summit of the apex were mitotically active and contributed to the developing shoot and floral structures. Thus, cells in L2 at the summit of vegetative apices are valid targets for transformation leading subsequently to modified germ line cells. Dissections to expose apices for DNA delivery inhibited mitotic activity; recovery periods greater than 48 h would be needed for restoration of normal activity. This suggests that a period of recovery from dissection would be beneficial for attempts at integrative transformation of apical cells.