Cellular basis of developmental plasticity observed in heterophyllous leaf formation of Ludwigia arcuata (Onagraceae)

When heterophyllous plants of Ludwigia arcuata Walt. (Onagraceae) were transferred from aerial condition to submergence, young developing leaves were matured into leaves with intermediate shape between aerial-type and submerged-type, showing spatulate shape (spoon-shaped). This change was also induced by the exposure of plants to ethylene. On the other hand, when the plants were transferred from submergence to aerial conditions, young developing leaves were matured into intermediate-type leaves with elliptic shape (spearhead shape). Anatomical analysis revealed that the formation of spatulate leaf was caused by the reduction of the number of epidermal cells aligned in the leaf transverse direction in the basal region of the leaf while the tip regions remained as before and did not respond to this treatment. During development, the ethylene-induced spatulate leaves showed that three types of alterations in epidermal cell division were involved in this process. Changes in the distribution of cell divisions in leaf lamina were detected by the first day of ethylene exposure, and changes in the orientation of cell division planes were detected by the second day. However, changes in the number of cells aligned in the leaf transverse direction were not detected by this time. Three days after ethylene exposure, frequency of cell divisions changed, and by the time changes of cell numbers aligned in the leaf transverse direction were observed. Thus, the formation of intermediate-type leaves in L. arcuata was ascribed to the alterations of cell division patterns which was induced by ethylene.     

DEVELOPMENTAL ANATOMY OF DIRECT SHOOT ORGANOGENESIS FROM LEAF PETIOLES OF VITIS VINIFERA (VITACEAE)

The anatomy of direct shoot organogenesis from leaf petioles of Vitis vinifera cv. French Colombard cultured in vitro was studied by light microscopy. Regenerating petiole stubs were fixed at 2- or 3-day intervals and sectioned longitudinally. By day 3 on regeneration medium, new cell divisions were observed. After 6 days, three distinct regions of meristematic activity were apparent within the expanding petiole stub: the wound-response, organogenic, and vascularization regions. In the organogenic region, rapid periclinal divisions of vacuolate outer cortical cells formed nodular bumps, many of which developed vascular strands and marginal meristems and formed adventitious leaves. Promeristems with small, densely staining cells and a distinct tunica layer also originated in the organogenic region, by cell division in the epidermal and subepidermal cell layers. With vascularization and the formation of leaf primordia, many promeristems became adventitious shoot meristems. Adventitious leaves and promeristems were initiated continuously from day 10 until day 33. Promeristems were often initiated near or upon adventitious leaves but could form either before or after the adventitious leaf developed. Adventitious leaves and shoot meristems developed vascular connections with the vascular bundles of the original expiant. The implication of this pattern of regeneration for Agrobacterium-mediated transformation of Vitis is discussed.     

HYBRIDIZATION,POLYPLOIDY AND ADVENTITIOUS GROWTH IN THE GENUS ASPLENIUM

Gametophytes of Asplenium platyneuron, A. rhizophyllum and A. monlanum were grown from spores under controlled conditions. Factors inhibiting and promoting germination and growth were determined. Leaves cut from sexually produced sporophytes of A. platyneuron, A. rhizophyllum and A. montanum were cultured under controlled conditions on agar slants. The leaves produced neoplastic growths of two kinds: (1) two-dimensional growths, (2) three-dimensional growths. The former developed into normal-appearing, rhizoid- and gametangia-producing, heart-shaped gametophytes having a diploid chromosome complement. The formation of two-dimensional growths was favored by conditions drastically reducing the energy supply of the initiating sporophytic cells and by destroying the correlation of the leaf. Three-dimensional growths most often developed into sporophytes unless conditions were limiting; in the latter case they transformed into two-dimensional growths. Both two- and three-dimensional growths developed from fragmented leaves excised from buds growing near the tips of acuminate fronds of A. ebenoides. The high frequency of two-dimensional adventitious growth leads to the conclusion that apospory may provide a mechanism for the production of fertile allotetraploids, by the fusion of diploid gametes of adventitiously produced diploid gametophytes, in the genus Asplenium and perhaps other fern genera, in contrast to that which has been previously suggested in the literature.     

Structure and development of the secretory cavities of Myrtus communis leaves

The structure and development of Myrtus communis L. secretory cavities has been studied in young and expanded leaves, using light and scanning electron microscope. Secretory cavities are continuously formed during leaf development, but in mature leaves the rhythm of their appearance shows steep decrease. Each secretory cavity is developed from a single epidermal cell, which undergoes a periclinal division followed by anticlinal and several oblique cell divisions. The lumen of the secretory cavity is initiated by cell wall separation, i.e., schizogenously. The secretory cells line the cavity, where the secreted material is collected. Secretory cavities are covered by modified epidermal cells, which do not seem to form any special aperture. Essential oils seem to be discharged after mechanical treatment of the leaf.     

THE DEVELOPMENTAL BASIS OF ANISOPHYLLY IN SELAGINELLA MARTENSII. I. INITIATION AND MORPHOLOGY OF GROWTH

The dorsiventral shoot system of Selaginella martensii is characterized by opposite pairs of ventral and dorsal leaves that are dimorphic in size and form. This study was undertaken to determine if the smaller dorsal leaf can be appropriately regarded as an arrested form of the larger leaf. Although the pattern of cell divisions and cell enlargement associated with leaf initiation is similar for both leaf types, the extent of localized growth results in distinctly larger primordia on the ventral side of the shoot. Ventral leaf primordia are also distinguished by the early formation of more extensive mesophyll tissue. Regression analysis of quantitative data on leaf length vs. position and leaf width vs. length indicates that the growth pattern of ventral and dorsal leaves is significantly different. These observations indicate that the developmental pathways of the dimorphic leaves of Selaginella martensii do not diverge at a relatively late developmental stage, but rather can be distinguished from inception.     

Apospory in leaf culture of staghorn fern (Platycerium bifurcatum)

The induction, origin, morphology, and ploidy of aposporous gametophytes produced on juvenile leaves of the fern Platycerium bifurcatum (Cav.) C. Chr. were studied. Leaf explants were grown on modified Murashige and Skoog medium with 0%, 0.01%, 0.1%, 1%, or 2% sucrose. A low sucrose concentration (0.01%) and wounding of the adaxial side of the leaf significantly increased the induction of aposporous gametophytes (90% of leaves produced gametophytes). Regeneration began as a proliferation of mainly epidermal cells on both sides of the leaf; subsequent development was similar to that shown by gametophytes originating from spores. Flow cytometric analysis of sporophytes and aposporous gametophytes revealed that both forms had the same ploidy level. On the basis of these findings, we propose a set of conditions which regularly and reproducibly induces apospory on most of the leaf explants of the fern P. bifurcatum.     

STRUCTURAL INVESTIGATIONS OF ASEXUAL REPRODUCTION IN NEPHROLEPIS EXALTATA AND PLATYCERIUM BIFURCATUM

Nephrolepis exaltata cv. Bostoniensis, the Boston fern, exhibits extreme stem dimorphism. The plant has orthotropic, dictyostelic shoots which bear pinnatifid leaves and plagiotropic, protostelic stolons which are aphyllous. Vegetative reproduction occurs by budding from primary and secondary stolons. Secondary stolons arise exogenously from derivatives of the apical cell of the primary stolon, whereas root primordia develop endogenously. Shoots develop in vivo when a creeping stolon makes contact with the substrate via extensive root proliferation. When stolon segments are excised and grown in vitro, secondary stolon primordia expand and initiate leaf primordia, forming new leafy shoots. In Platycerium bifurcatum, the staghorn fern, asexual propagation occurs on ageotropic roots ramifying among the basal nest fronds. Root bud initiation is marked by root tip hypertrophy following cortical parenchyma expansion. Root apical cell derivatives produce the bud apex; the root apical cell remains separate from the developing root bud. Superficially, vegetative reproduction in Nephrolepis and Platycerium appears to involve unusual organs. However, both ferns exhibit leafy bud development from distinct sites of origin, not from undetermined primordia or from direct transformation of root to shoot. Thus, distinctness of organ types is maintained in these two ferns and no evidence for interconvertibility of organ types has been found.     

The effect of sucrose on the differentiation of excised fern leaf tissue into either gametophytes of sporophytes

Excised juvenile leaves of Microgramma vacciniifolia (Polypodiaceae) develop sporophytic regenerants when grown on mineral agar with sucrose. The ratio of sporophytes to gametophytes produced from the leaf tissue increases with higher percentages of sucrose such that at 4% sucrose, the induction of aposporous gametophytes is a rare occurrence. Experiments varying the osmotic potential with sorbitol and those holding the osmotic potential of the culture medium constant while varying the sucrose level indicate that the effect of sucrose on the differentiation of fern leaf tissue into either gametophyte or sporophyte is nutritional rather than osmotic. A significant effect of sucrose in altering the differentiation of fern leaf tissue is the increased rate of senescence promoted by high sucrose concentrations.     

A DEVELOPMENTAL STUDY OF SILICIFICATION IN THE ABAXIAL EPIDERMAL CELLS OF SUGARCANE LEAF BLADES USING SCANNING ELECTRON MICROSCOPY AND ENERGY DISPERSIVE X-RAY ANALYSIS

A developmental study of the accumulation of silicon and other elements in the abaxial epidermis of sugarcane (Saccharum officinarum L.) leaf blades using scanning electron microscopy and energy dispersive x-ray analysis showed that accumulation of silicon progresses at different rates in each epidermal cell type. In basal cells of two-celled microhairs and in prickles there is accumulation of silicon while the leaf is immature and still enclosed within the spindle cluster of leaves and not involved in transpiration. After transpiration begins, all epidermal cells rapidly accumulate silicon. However, there are differences in the rate of silicon accumulation and in the maximum amount of silicon accumulation among the various cell types. This may relate to differences in their physiology or structure.     

Cellular basis for dimethipin-induced loss of leaf turgor and desiccation

Dimethipin-induced increase in transpiration from kidney bean leaves (Phaseolus vulgaris L. cv. Black Valentine) was not correlated with stomatal aperture. From analysis of the kinetics of water loss from excised kidney bean leaves, it was concluded that the increase in transpiration was due almost entirely to an increase in the cuticular component. Both light and scanning electron microscopic analysis of dimethipin-treated leaves indicated that the first cells to be affected by dimethipin were the epidermal cells. These results suggest that dimethipin causes a loss of leaf turgor and desiccation by disrupting epidermal cells, thereby removing a major barrier for water loss from the leaf.Mention of trademark or proprietary product does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable.     

QUANTITATIVE STUDIES OF THE VEGETATIVE SHOOT APEX OF EQUISETUM SCIRPOIDES

Equisetum scirpoides Michx., propagated from a single clone, was grown in a controlled growth chamber at 24 ± 1 C under a photoperiod of 16 hr light/8 hr darkness. The apical cell of aerial vegetative shoots gives rise to derivatives (merophytes) in a helical sequence. Each newly formed merophyte divides anticlinally to form two superposed cells that are parallel to a lateral face of the apical cell. Radial longitudinal divisions then take place in the two superposed cells. Shoot tips were fixed every 2 hr for 24 hr to determine the mitotic index of the apical cell, six subjacent cells, and the remaining cells above the level of leaf initiation. Average mitotic indices for the 24-hr period were 3.9%, 3.9%, and 7.0%, respectively. The results indicate that the apical cell is quite active mitotically; there was no clear evidence of endopolyploidy in cells of the shoot apex, young leaves or in the developing cortex, based upon cytophotometric measurements of DNA content.     

MORPHOLOGY OF MARSILEA VESTITA. I. ONTOGENY AND MORPHOLOGY OF THE SUBMERGED AND LAND FORMS OF THE JUVENILE LEAVES

During their ontogeny, the primordia of the juvenile leaves of Marsilea plants in sterile culture develop 1, 2 or 4 marginal meristems, and these, in turn, contribute cells to the young leaf by anti- and periclinal cell divisions. The final leaves are unifid, bifid, or quadrifid, depending on how many marginal meristems develop, and this is determined early in the ontogeny of the leaf. The mechanism which determines whether or not a marginal meristem develops may fluctuate, as shown by the existence of trifid leaves. Two forms of juvenile leaves are produced, those in a liquid medium, which in many respects resemble the adult quadrifid submerged leaves, and those on a solid medium, which in many respects resemble the adult land leaves.     

Histological changes associated with acquisition of competence for shoot regeneration in leaf discs ofSaintpaufla ionantha xconfusa hybrid (African violet) cultured in vitro

In leaf discs ofSaintpaulia ionantha xconfusa hybrid (cv. Virginia) cultured on shoot-inducing medium, periclinal divisions were initiated in epidermal cells 3–5 days after explant isolation. This timing coincided with the time for competence acquisition determined in tissue-transfer experiments. Some of the daughter cells from periclinal divisions formed the target cells which divided both anticlinally and periclinally to form cell division centers (meristemoids), precursors of adventitious shoots. The target cells were not morphologically distinct from other epidermal cells at the light microscope level. It is suggested that the periclinal divisions in epidermal cells represent the dedifferentiation phase during which target (competent) cells are formed. Once the cells have acquired the ability to divide periclinally, both dedifferentiation and shoot induction occur in the presence of exogenous plant hormones.Abbreviations SIM Shoot-inducing medium     

Regulation of a Sub-sexual Life Cycle in a Moss: Evidence for the Occurrence of a Factor for Apogamy in Physcomitrium

As demonstrated in our earlier studies, the differentiationof apogamous sporophytes on the secondary protonema of the mossPhyscomitrium pyriforme Brid. requires an exogenous supply ofsucrose in the medium. In the present work, similar differentiationwas observed in the leaf cells of aged gametophytes. The experimentsindicate an accumulation in the leaves of a sporophytic factorwhich initiates a de novo differentiation of sporophytes fromleaf cells without the intervention of sexual reproduction.In the absence of sucrose, the factor for apogamy was not present.Highlight intensity (5000–6000 lx) also inhibited itsproduction. There was no evidence that its presence interferedwith or inhibited production of gameto-phores. Growth regulatorssuch as IAA and kinetin altered only the effectiveness of thissporophytic factor, demonstrating that it was endogenous. Sporogenesisin the apogamous sporophytes took place without orthodox meiosis. Results obtained by using different exogenous environments forthe in vitro differentiation of callus into gametophytes orsporophytes are also reported. These support our contentionthat there is an accumulation of a sporophytic factor in thegametophytic callus cells, which is diluted during the processof differentiation. The morpho-regulatory influence of lightin the differentiation of apical cells with three cutting facesfrom unorganized callus is also considered.     

LIRIODENDRON (MAGNOLIACEAE) FROM THE MIOCENE CLARKIA FLORA OF IDAHO

Miocene Liriodendron carpels, whole fruiting structures and leaves from Clarkia and Oviatt Creek sites in northern Idaho are preserved as imprints and compressed fossils in soft lacustrine clays. The isolated carpels are indistinguishable from those described as L. hesperia Berry from the Spokane Latah flora. Fruit aggregates from the type Clarkia and Oviatt Creek localities and leaves from three Clarkia sites are considered to be within the range of variation of the single species L. hesperia. Comparisons were made regarding leaf architecture, lower leaf epidermal structures, leaf flavonoid and steroid analysis, morphological features of receptacles and carpels, and the venation pattern of carpels of the fossil material to the two extant species, L. tulipifera L. (native to southeastern United States) and L. chinense Sarg. (native to southeastern Asia). Leaf architecture features analyzed by standard statistical and canonical tests and marginal venation patterns near the base of leaves suggest that L. hesperia is more similar to L. tulipifera, whereas the size dimensions of lower epidermal cells and the common presence of two sterane compounds imply that L. hesperia is more similar to L. chinense. The fossil species, however, is a distinct taxon indicated by statistical discriminant and canonical tests, leaf base shape, often smaller epidermal cell dimensions, and the shape of round receptacle carpel scars. Both the fossil and the two living Liriodendron species are associated with comparable mixed mesophytic floras.     

Morphogenetic role of kinetin and abscisic acid in the moss Physcomitrium

Summary In the presence of kinetin, a supposedly gametophytic bud inducing substance, the secondary protonema of the moss Physcomitrium pyriforme Brid., as well as producing leafy gametophytes, continued to exhibit its normal tendency of forming sporophytic buds (i.e. buds with apical cells having two cutting faces). Also remarkable was that callus derived from the secondary protonema, when cultured in a kinetin supplemented liquid medium, formed exclusively apogamous sporophytic buds with a virtual exclusion of gametophytes. In the presence of abscisic acid, the elongation of protonemal cells as well as their differentiation was markedly suppressed. This effect was manifest even when abscisic acid was used in conjunction with kinetin. It is suggested that rather than having a morphoregulatory role, kinetin may be responsible merely for enhancing cell proliferation. The determination of an apical cell with two cutting faces (sporophytic) or one with three cutting faces (gametophytic) is under the control of other factors both external, (e.g. sucrose) and internal. It is proposed that abscisic acid can suppress the usual differentiational capacity of the moss tissue, even in a favourable environment.     

Developmental process of sun and shade leaves in Chenopodium album L.

The authors’ previous study of Chenopodium album L. revealed that the light signal for anatomical differentiation of sun and shade leaves is sensed by mature leaves, not by developing leaves. They suggested that the two‐cell‐layered palisade tissue of the sun leaves would be formed without a change in the total palisade tissue cell number. To verify that suggestion, a detailed study was made of the developmental processes of the sun and shade leaves of C. album with respect to the division of palisade tissue cells (PCs) and the data was expressed against developmental time (leaf plastochron index, LPI). The total number of PCs per leaf did not differ between the sun and shade leaves throughout leaf development (from LPI ?1 to 10). In both sun and shade leaves, anticlinal cell division of PCs occurred most frequently from LPI ?1 to 2. In sun leaves, periclinal division of PCs occurred synchronously with anticlinal division. The constancy of the total number of PCs indicates that periclinal divisions occur at the expense of anticlinal divisions. These results support the above suggestion that two‐cell‐layered palisade tissue is formed by a change of cell division direction without a change in the total number of PCs. PCs would be able to recognize the polarity or axis that is perpendicular to the leaf plane and thereby change the direction of their cell divisions in response to the light signal from mature leaves.     

FLEXIBILITY IN ONTOGENY AS SHOWN BY THE CONTRIBUTION OF THE SHOOT APICAL LAYERS TO LEAVES OF PERICLINAL CHIMERAS

The development of leaves on apically stable, periclinal chimeras was studied in a number of dicot genera. The mutant cell layers of the shoot apex and the tissues derived from them were as active developmentally as the normal layers. Ontogeny was the same in these chimeras as in nonchimeras, and growth of their leaves can be outlined as follows. Formation of the buttress, the axis, and the lamina of simple dicot leaves were independent events. In each the first growth included derivatives of the apical layers, usually three in number, found in the apex of the shoot and the lateral buds. Most cell divisions in the outer layers (L-I and L-II) were anticlinal relative to the new structures. Therefore, in the proximal regions of the buttress, axis (petiole and midrib), and lamina, the derivative cells of L-I and L-II were usually present in single layers. The rest of the internal tissue was from L-III. As formation of the axis and the lamina proceeded, derivatives of L-II replaced L-III internally in the distal and marginal regions leaving cells of L-III behind. Both the determinate growth of leaves and the pattern of cell divisions at and near the leading edges of growth meant that no cells in the leaf were comparable to the initial cells of the shoot apex. As the lamina extended, there were extensive intercalary cell divisions, both anticlinal and periclinal, so that in any given region of a leaf the layers of internal cells were from either L-II or L-III. At any point along the axis, L-III participated or did not participate in laminar extension. At any given stage in laminar growth either of two sister cells in any internal layer divided either a few times or extensively. The extreme variability in direction and frequency of cell division during leaf development was under an overriding genetic control, which resulted in the normal or typical size, shape and thickness of leaves.     

Contributions of photosynthetic and non‐photosynthetic cell types to leaf respiration in Vicia faba L. and their responses to growth temperature

In intact leaves, mitochondrial populations are highly heterogeneous among contrasting cell types; how such contrasting populations respond to sustained changes in the environment remains, however, unclear. Here, we examined respiratory rates, mitochondrial protein composition and response to growth temperature in photosynthetic (mesophyll) and non‐photosynthetic (epidermal) cells from fully expanded leaves of warm‐developed (WD) and cold‐developed (CD) broad bean (Vicia faba L.). Rates of respiration were significantly higher in mesophyll cell protoplasts (MCPs) than epidermal cell protoplasts (ECPs), with both protoplast types exhibiting capacity for cytochrome and alternative oxidase activity. Compared with ECPs, MCPs contained greater relative quantities of porin, suggesting higher mitochondrial surface area in mesophyll cells. Nevertheless, the relative quantities of respiratory proteins (normalized to porin) were similar in MCPs and ECPs, suggesting that ECPs have lower numbers of mitochondria yet similar protein complement to MCP mitochondria (albeit with lower abundance serine hydroxymethyltransferase). Several mitochondrial proteins (both non‐photorespiratory and photorespiratory) exhibited an increased abundance in response to cold in both protoplast types. Based on estimates of individual protoplast respiration rates, combined with leaf cell abundance data, epidermal cells make a small but significant (2%) contribution to overall leaf respiration which increases twofold in the cold. Taken together, our data highlight the heterogeneous nature of mitochondrial populations in leaves, both among contrasting cell types and in how those populations respond to growth temperature.